science/medicine

Adequate humidification in long-term jet ventilation is a critical aspect in terms of clinical safety. To assess a prototype of an electronic jet-ventilator and its humidification system. Forty patients with respiratory insufficiency were randomly allocated to one of four groups. The criterion for inclusion in this study was respiratory insufficiency exhibiting a Murray score above 2. The four groups of patients were ventilated with three different respirators and four different humidification systems. Patients in groups A and B received superimposed high-frequency jet ventilation (SHFJV) by an electronic jet-ventilator either with (group A) or without (group B) an additional humidification system. Patients in group C received high-frequency percussive ventilation (HFPV) by a pneumatic high-frequency respirator, using a hot water humidifier for warming and moistening the inspiration gas. Patients in group D received conventional mechanical ventilation using a standard intensive care unit respirator with a standard humidification system. SHFJV and HFPV were used for a period of 100 h (4days). < 0.05) but rose to an average of 98 ± 2.8% after 2 h. The average percentage across all four groups amounted to 98 ± 0.4% after 2 h. Inflammation of the tracheal mucosa was found in patients in group B and the mucosal injury score (MIS) was significantly higher than in all the other groups. Patients in groups A, C and D showed no severe evidence of airway damage, exhibiting adequate values of relative humidity and temperature of the inspired gas. The problems of humidification associated with jet ventilation can be fully prevented by using this new jet-ventilator. These data were sustained by nondeteriorating MIS values at the end of the 4-day study period in groups A, C and D.

In eukaryotic cells, proteins are translocated across the ER membrane through a continuous ribosome-translocon channel. It is unclear to what extent proteins can fold already within the ribosome-translocon channel, and previous studies suggest that only a limited degree of folding (such as the formation of isolated α-helices) may be possible within the ribosome. We have previously shown that the conformation of nascent polypeptide chains in transit through the ribosome-translocon complex can be probed by measuring the number of residues required to span the distance between the ribosomal P-site and the lumenally disposed active site of the oligosaccharyl transferase enzyme (J. Biol. Chem 271: 6241-6244).Using this approach, we now show that model segments composed of residues with strong helix-forming properties in water (Ala, Leu) have a more compact conformation in the ribosome-translocon channel than model segments composed of residues with weak helix-forming potential (Val, Pro). The main conclusions from the work reported here are (i) that the propensity to form an extended or more compact (possibly α-helical) conformation in the ribosome-translocon channel does not depend on whether or not the model segment has stop-transfer function, but rather seems to reflect the helical propensities of the amino acids as measured in an aqueous environment, and (ii) that stop-transfer sequences may adopt a helical structure and integrate into the ER membrane at different times relative to the time of glycan addition to nearby upstream glycosylation acceptor sites.

Previous studies by Volloch and coworkers have reported that during the expression of high levels of β-globin mRNA in the spleen of anemic mice, they could also detect small but significant levels of an antisense (AS) globin RNA species, which they postulated might have somehow arisen by RNA-directed RNA synthesis. For two reasons we undertook to confirm and possibly extend these studies. First, previous studies in our lab have focussed on what is an unequivocal example of host RNA-directed RNA polymerase activity on the RNA genome of human hepatitis delta virus. Second, if AS globin species do exist they could in turn form double-stranded RNA species which might induce post-transcriptional gene silencing, a phenomenon somehow provoked in eukaryotic cells by AS RNA sequences. We reexamined critical aspects of the previous globin studies. We used intraperitoneal injections of phenylhydrazine to induce anemia in mice, as demonstrated by the appearance and ultimate disappearance of splenomegaly. While a 30-fold increase in globin mRNA was detected in the spleen, the relative amount of putative AS RNA could be no more than 0.004%. Contrary to earlier reports, induction of a major increase in globin transcripts in the mouse spleen was not associated with a detectable level of antisense RNA to globin mRNA.

Serum and synovial tissue expression of the matrix metalloproteinase (MMP)-2 and -9 and their molecular regulators, MMP-14 and TIMP-2 was examined in 28 patients with inflammatory early synovitis and 4 healthy volunteers and correlated with the presence of erosions in the patients. Immunohistological staining of MMP-2, MMP-14 and TIMP-2 localized to corresponding areas in the synovial lining layer and was almost absent in normal synovium. Patients with radiographic erosions had significantly higher levels of active MMP-2 than patients with no erosions, suggesting that activated MMP-2 levels in synovial tissue may be a marker for a more aggressive synovial lesion. In cancer the gelatinases [matrix metalloproteinase (MMP)-2 and MMP-9] have been shown to be associated with tissue invasion and metastatic disease. In patients with inflammatory arthritis the gelatinases are expressed in the synovial membrane, and have been implicated in synovial tissue invasion into adjacent cartilage and bone. It is hypothesized that an imbalance between the activators and inhibitors of the gelatinases results in higher levels of activity, enhanced local proteolysis, and bone erosion. To determine whether the expression and activity levels of MMP-2 and MMP-9, and their regulators MMP-14 and tissue inhibitor of metalloproteinase (TIMP), are associated with early erosion formation in patients with synovitis of recent onset. A subset of 66 patients was selected from a larger early synovitis cohort on the basis of tissue availability for the study of synovial tissue and serum gelatinase expression. Patients with peripheral joint synovitis of less than 1 years' duration were evaluated clinically and serologically on four visits over a period of 12 months. At the initial visit, patients underwent a synovial tissue biopsy of one swollen joint, and patients had radiographic evaluation of hands and feet initially and at 1year. Serum MMP-1, MMP-2, MMP-9, MMP-14, and TIMP-1 and TIMP-2 levels were determined, and synovial tissue was examined by immunohistology for the expression of MMP-2 and MMP-9, and their molecular regulators. Gelatinolytic activity for MMP-2 and MMP-9 was quantified using a sensitive, tissue-based gel zymography technique. Four healthy individuals underwent closed synovial biopsy and their synovial tissues were similarly analyzed. ). Tissue expression of MMP-2 and MMP-9, however, did not correlate with the serum levels of these enzymes. < 0.001). MMP-2 and MMP-9 are thought to play an important role in the evolution of joint erosions in patients with an inflammatory arthritis. Most studies have concentrated on the contribution of MMP-9 to the synovitis, because synovial fluid and serum MMP-9 levels are markedly increased in inflammatory arthropathies. Previously reported serum levels of MMP-9 have varied widely. In the present sample of patients with synovitis of recent onset, serum MMP-9 levels were elevated in only 21%. Moreover, these elevations were not specific for RA, the tissue expression of MMP-9 was focal, and the levels of MMP-9 activity were not well correlated with early erosions. Although serum MMP-2 levels were not of prognostic value, high synovial tissue levels of MMP-2 activity were significantly correlated with the presence of early erosions. This may reflect augmented activation of MMP-2 by the relatively high levels of MMP-14 and low levels of TIMP-2 seen in these tissues. We were able to localize the components of this trimolecular complex to the synovial lining layer in consecutive tissue sections, a finding that is consistent with their colocalization. In conclusion, we have provided evidence that active MMP-2 complexes are detectable in the inflamed RA synovium and may be involved in the development of early bony erosions. These results suggest that strategies to inhibit the activation of MMP-2 may have the potential for retarding or preventing early erosions in patients with inflammatory arthritis.

We analyzed the causes and results of utilization of critical care services in the special care unit in patients after surgical procedures performed by the hepatobiliary surgical service during a 23-month period. = 0.041). Respiratory failure was the predominant component of all complications after hepatobiliary surgery. No clinically useful predictors of eventual outcome could be identified.

. The objective is to explore the role of the aquatic reservoir on the persistence of endemic cholera as well as to define minimum conditions for the development of epidemic and endemic cholera. The reproduction rate of cholera in a community is defined by the product of social and environmental factors. The importance of the aquatic reservoir depends on the sanitary conditions of the community. Seasonal variations of contact rates force a cyclical pattern of cholera outbreaks, as observed in some cholera-endemic communities. infection in endemic populations as well as a better description of the relationship between dose and virulence.

Data provided by the social sciences as well as genetic research suggest that the 8-10 million Roma (Gypsies) who live in Europe today are best described as a conglomerate of genetically isolated founder populations. The relationship between the traditional social structure observed by the Roma, where the Group is the primary unit, and the boundaries, demographic history and biological relatedness of the diverse founder populations appears complex and has not been addressed by population genetic studies. Recent medical genetic research has identified a number of novel, or previously known but rare conditions, caused by private founder mutations. A summary of the findings, provided in this review, should assist diagnosis and counselling in affected families, and promote future collaborative research. The available incomplete epidemiological data suggest a non-random distribution of disease-causing mutations among Romani groups. Although far from systematic, the published information indicates that medical genetics has an important role to play in improving the health of this underprivileged and forgotten people of Europe. Reported carrier rates for some Mendelian disorders are in the range of 5 -15%, sufficient to justify newborn screening and early treatment, or community-based education and carrier testing programs for disorders where no therapy is currently available. To be most productive, future studies of the epidemiology of single gene disorders should take social organisation and cultural anthropology into consideration, thus allowing the targeting of public health programs and contributing to the understanding of population structure and demographic history of the Roma.

This study describes the upregulation of neurokinin 1 and bradykinin 2 receptors in dorsal root ganglion (DRG) neurons in the course of antigen-induced arthritis (AIA) in the rat knee. In the acute phase of AIA, which was characterized by pronounced hyperalgesia, there was a substantial bilateral increase in the proportion of lumbar DRG neurons that express neurokinin 1 receptors (activated by substance P) and bradykinin 2 receptors. In the chronic phase the upregulation of bradykinin 2 receptors persisted on the side of inflammation. The increase in the receptor expression is relevant for the generation of acute and chronic inflammatory pain. Ongoing pain and hyperalgesia (enhanced pain response to stimulation of the tissue) are major symptoms of arthritis. Arthritic pain results from the activation and sensitization of primary afferent nociceptive nerve fibres ('pain fibres') supplying the tissue (peripheral sensitization) and from the activation and sensitization of nociceptive neurons in the central nervous system (central sensitization). After sensitization, nociceptive neurons respond more strongly to mechanical and thermal stimulation of the tissue, and their activation threshold is lowered. The activation and sensitization of primary afferent fibres results from the action of inflammatory mediators such as bradykinin (BK), prostaglandins and others on membrane receptors located on these neurons. BK is a potent pain-producing substance that is contained in inflammatory exudates. Up to 50% of the primary afferent nerve fibres have receptors for BK. When primary afferent nerve fibres are activated they can release neuropeptides such as substance P (SP) and calcitonin gene-related peptide from their sensory endings in the tissue. SP contributes to the inflammatory changes in the innervated tissue (neurogenic inflammation), and it might also support the sensitization of nociceptive nerve fibres by binding to neurokinin 1 (NK1) receptors. NK1 receptors are normally expressed on a small proportion of the primary afferent nerve fibres. Because the expression of receptors on the primary afferent neurons is essential for the pain-producing action of inflammatory mediators and neuropeptides, we investigated in the present study whether the expression of BK and NK1 receptors on primary afferent neurons is altered during the acute and chronic phases of an antigen-induced arthritis (AIA). AIA resembles in many aspects the inflammatory process of human rheumatoid arthritis. Because peptide receptors are expressed not only in the terminals of the primary afferent units but also in the cell bodies, we removed dorsal root ganglia (DRGs) of both sides from control rats and from rats with the acute or chronic phase of AIA and determined, after short-term culture of the neurons, the proportion of DRG neurons that expressed the receptors in the different phases of AIA. We also characterized the inflammatory process and the nociceptive behaviour of the rats in the course of AIA. . After immunization, m-BSA was injected into the right knee joint cavity to induce arthritis. The joint swelling was measured at regular intervals. Nociceptive (pain) responses to mechanical stimulation of the injected and the contralateral knee were monitored in the course of AIA. Groups of rats were killed at different time points after the induction of AIA, and inflammation and destruction in the knee joint were graded by histological examination. The DRGs of both sides were dissected from segments L1–L5 and C1–C7 from arthritic rats, from eight immunized rats without arthritis and from ten normal control rats. Excised DRGs were dissociated into single cells which were cultured for 18 h. )-Lys-BK. ). The initial phase of AIA was characterized by strong joint swelling and a predominantly granulocytic infiltration of the synovial membrane and the joint cavity (acute inflammatory changes). In the later phases of AIA (10–84 days after induction of AIA) the joint showed persistent swelling, and signs of chronic arthritic alterations such as infiltration of mononuclear leucocytes, hyperplasia of synovial lining layer (pannus formation) and erosions of cartilage and bone were predominant. The contralateral knee joints appeared normal at all time points. Destruction was observed only in the injected knee but some proteoglycan loss was also noted in the non-injected, contralateral knee. In the acute and initial chronic phases of AIA (1–29 days) the rats showed mechanical hyperalgesia in the inflamed knee (limping, withdrawal response to gentle pressure onto the knee). In the acute phase (up to 9 days) a pain response was also seen when gentle pressure was applied to the contralateral knee. ]–SP in three experiments, showing that SP–gold was bound to NK1 receptors. = 5) of AIA was the binding of BK–gold decreased by the coadministration of BK–gold and the B1 agonist. By contrast, in these experimental groups the binding of BK–gold was suppressed by the coadministration of the B2 agonist. These results show that B2 receptors, but not B1 receptors, were expressed in both normal animals and in animals with AIA. These results show that in AIA in the rat the expression of SP-binding and BK-binding sites in the perikarya of DRGs L1–L5 is markedly upregulated in the course of knee inflammation. Although the inflammation was induced on one side only, the initial changes in the binding sites were found in the lumbar DRGs of both sides. No upregulation of SP-binding or BK-binding sites was observed in the cervical DRGs. The expression of SP-binding sites was upregulated only in the first days of AIA, that is, in the acute phase, in which the pain responses to mechanical stimulation were most pronounced. By contrast, the upregulation of BK-binding sites on the side of AIA persisted for up to 42 days, that is, in the acute and chronic phase of AIA. Only the B2 receptor, not the B1 receptor, was upregulated. The coincidence of the enhanced expression of NK1 and BK receptors on sensory neurons and the pain behaviour suggests that the upregulation of these receptors is relevant for the generation and maintenance of arthritic pain. In the acute phase of AIA, approximately 50% of the lumbar DRG neurons showed an expression of SP-binding sites. Because peptide receptors are transported to the periphery, the marked upregulation of SP-binding receptors probably leads to an enhanced density of receptors in the sensory endings of the primary afferent units. This will permit SP to sensitize more neurons under inflammatory conditions than under normal conditions. However, the expression of NK1 receptors was upregulated only in the acute phase of inflammation, suggesting that SP and NK1 receptors are less important for the generation of hyperalgesia in the chronic phase of AIA. Because BK is one of the most potent algesic compounds, the functional consequence of the upregulation of BK receptors is likely to be of immediate importance for the generation and maintenance of inflammatory pain. The persistence of the upregulation of BK receptors on the side of inflammation suggests that BK receptors should be an interesting target for pain treatment in the acute and chronic phases. Only B2 receptors were identified in normal animals and in rats with AIA. This is surprising because previous pharmacological studies have provided evidence that, during inflammation, B1 receptors can be newly expressed. Receptor upregulation in the acute phase of AIA was bilateral and almost symmetrical. However, hyperalgesia was much more pronounced on the inflamed side. It is most likely that receptors on the contralateral side were not readily activated because in the absence of gross inflammation the local concentration of the ligands BK and SP was probably quite low. We hypothesize that the bilateral changes in receptor expression are generated at least in part by mechanisms involving the nervous system. Symmetrical segmental changes can be produced only by the symmetrical innervation, involving either the sympathetic nervous system or the primary afferent fibres. Under inflammatory conditions, primary afferent fibres can be antidromically activated bilaterally in the entry zone of afferent fibres in the spinal cord, and it was proposed that this antidromic activation might release neuropeptides and thus contribute to neurogenic inflammation. Because both sympathetic efferent fibres and primary afferent nerve fibres can aggravate inflammatory symptoms, it is also conceivable that they are involved in the regulation of receptor expression in primary afferent neurons. A neurogenic mechanism might also have been responsible for the bilateral degradation of articular cartilage in the present study.

) ratios were statistically significantly higher in the dexmedetomidine group. Dexmedetomidine provides important postsurgical analgesia and appears to have no clinically important adverse effects on respiration in the surgical patient who requires intensive care. ]. Cardiovascular stability was demonstrated, with significant reductions in rate-pressure product during sedation and over the extubation period. -mediated reduction in sympathetic tone. Therefore, it should be possible to continue sedation with dexmedetomidine over the stressful extubation period without concerns over respiratory depression, while ensuring that haemodynamic stability is preserved. ]) after extubation in the ICU. Patients who participated in the present study were admitted after surgery to our general or cardiothoracic ICUs, and were expected to receive at least 6 h of postsurgical sedation and artificial ventilation. ] of 3 or greater while the patients were intubated, and infusions of study drug were continued for a maximum of 6 h after extubation to achieve a Ramsay Sedation Score of 2 or greater. The patients were intubated and ventilated with oxygen-enriched air to attain acceptable arterial blood gases, and extubation occurred when clinically indicated. All patients received supplemental oxygen after extubation, which was delivered by a fixed performance device. Assessment of pain was by direct communication with the patient. comparisons. Of the 40 patients who participated in the study, seven patients could not be included in the analysis of respiratory function because they did not receive a study drug infusion after extubation. Consequently, data from 33 patients are used in the analysis of respiratory function; 16 received dexmedetomidine and 17 placebo. Inadequate arterial blood gas analysis was available in five patients (two from the dexmedetomidine group, and three from the placebo group). There were no significant differences in patient characteristics and operative details between the groups. = 0.040). ) for the 6 h after extubation. ). There were no adverse respiratory events seen in either the dexmedetomidine or placebo group. Respiratory rate for the 6-h periods before and after extubation. (Filled circle) Dexmedetomidine; (Empty circle) placebo. Values are expressed as mean ± standard deviation. ) for the 6-h periods before and after extubation, and baseline values (B) on admission to ICU immediately after surgery. (Filled circle) Dexmedetomidine; (Empty circle) placebo. Values are expressed as mean ± standard deviation. ratio for the 6-h periods before and after extubation, and baseline values (B) on admission to ICU immediately after surgery. (Filled circle) Dexmedetomidine; (Empty circle) placebo. Values are expressed as mean ± standard deviation. ]. were mildly decreased and increased, respectively. There was a rightward shift and depression of the hypercapnic response with infusions of 1.0 and 2.0 μg/kg. between the groups. We can not therefore determine from this study whether dexmedetomidine has any benefits over morphine from a respiratory perspective. analysis reveals that the data have 80% power to detect a significant difference (α value 0.05). Further studies are obviously required. ] that dexmedetomidine provides cardiovascular stability, with a reduction in rate-pressure product over the extubation period. A sedative agent that has analgesic properties, minimal effects on respiration and offers ischaemia protection would have enormous potential in the ICU. Dexmedetomidine may fulfill all of these roles, but at present we can only conclude that dexmedetomidine has no deleterious clinical effects on respiration when used in doses that are sufficient to provide adequate sedation and effective analgesia in the surgical population requiring intensive care.

Treatment of mouse F9 embryonal carcinoma cells with all-trans retinoic acid (T-RA) induces differentiation into primitive endodermal type cells. Differentiation requires the action of the receptors for all trans, and 9cis-retinoic acid (RAR and RXR, respectively) and is accompanied by growth inhibition, changes in cell morphology, increased apoptosis, proteolytic degradation of the RARγ2 receptor, and induction of target genes. 135 expression induces a novel differentiation pathway characterised by the appearance of cells with an atypical elongated morphology which are cAMP resistant. 135 is required for normal F9 cell differentiation. Hence, in addition to transactivators, targeted proteolysis of basal transcription factors also plays an important role in gene regulation in response to physiological stimuli.

A growing number of laboratories are using the mouse as a model system in developmental biology as well as in molecular biology. Surprisingly, most of these laboratories do not have reliable computerized systems to track these animals, and the few commercial solutions available are expensive. We thus developed MICE (Mouse Information and Classification Entity), a program aimed at facilitating the monitoring of animals in animal facilities. This program consists of a virtual facility in which scientists can perform all the tasks done in the real world (i.e., receiving animals, breeding them, preparing cage labels, etc.). Recording of each animal (birth date, cage number, ID number, tail analysis number, parents, genetic status, genetic background, etc.) enables reliable tracking. According to any parameter of interest, animals can then be identified, grouped, sorted, moved, and so forth. Crossings are automatically processed by the program. For example, new genetic backgrounds, generation number, and anticipated due dates are determined. The program also reminds the user when new births are expected and entering newborn animals only requires a few clicks. The genealogy of each animal can be determined in two different ways, one being the visualization of a genealogical tree from which information of ancestors can be retrieved. This standalone program, that will be distributed free of charge to academic laboratories requesting a license, represents a new and valuable tool for all animal facility users, and permits simple and reliable tracking and retrieving of animals.

The synovial membrane (SM) of affected joints in ankylosing spondylitis (AS) is infiltrated by germinal center-like aggregates (foci) of lymphocytes similar to rheumatoid arthritis (RA). We characterized the rearranged heavy chain variable segment (VH) genes in the SM for gene usage and the mutational pattern to elucidate the B lymphocyte involvement in AS. Cryosections from an AS-derived SM were stained for B and T lymphocytes. B cells were isolated from different areas of a focus. The rearranged VH genes were amplified by semi-nested polymerase chain reaction (PCR) using oligonucleotides specific for the six different VH families and heavy chain joining segments (JHs). PCR products were cloned and sequenced. Fifty-nine of 70 different heavy chain gene rearrangements were potentially functional. Most of the rearranged genes were mutated (range, 1–15%). Thirty of 70 products had a mutational pattern typical for antigen selection. Most of the rearranged VH genes belonged to the VH3 family (54%), consistent with data from healthy donors and patients with RA, while VH4 genes, in contrast to RA, were identified less frequently (10%) and VH5 genes were over-represented (11%). In contrast to RA, neither VH6 genes nor the autoimmunity-prone VH4-34 were seen, whereas another autoimmunity-prone gene, V3-23, was predominantly used (11%). One VH1-derived and one VH3-derived B cell clone were expanded. CDR3 were shorter and more variable in length than in RA. Comparable with RA and reactive arthritis, there is a biased repertoire of selected VH genes, whereas the panel of represented genes is different and less clonal expansion was observed.

Interest in methanogens from ruminants has resulted from the role of methane in global warming and from the fact that cattle typically lose 6 % of ingested energy as methane. Several species of methanogens have been isolated from ruminants. However they are difficult to culture, few have been consistently found in high numbers, and it is likely that major species of rumen methanogens are yet to be identified. , an organism not previously described in the rumen. The third cluster contained rDNA sequences that may form a novel group of rumen methanogens. The current set of 16S rRNA hybridization probes targeting methanogenic Archaea does not cover the phylogenetic diversity present in the rumen and possibly other gastro-intestinal tract environments. New probes and quantitative PCR assays are needed to determine the distribution of the newly identified methanogen clusters in rumen microbial communities.

, was cloned in 1998 and the encoded protein, cystinosin, was predicted to be a lysosomal membrane protein. . underlying cystinosis. Furthermore, our work has brought to light the existence of a differential pattern of expression between the human and murine homologues, providing critical information for the generation of a mouse model for cystinosis.

Head extension and excessive laryngoscope blade levering motion (LBLM) are undesirable during airway management of trauma patients. We hypothesized that laryngoscopy with a modified blade facilitating glottic exposure by balloon inflation would reduce head extension and LBLM. -tests. Laryngoscopic view grade and oxygen saturation were also determined. <0.001). Laryngoscopic view was approximately identical with both blades, and oxygen saturation was always above 97%. Balloon laryngoscopy reduces head extension and LBLM under simulated cervical spine precautions.

Axillary node status after induction chemotherapy for locally advanced breast cancer has been shown on multivariate analysis to be an independent predictor of relapse. However, it has been postulated that responders to induction chemotherapy with a clinically negative axilla could be spared the burden of lymphadenectomy, because most of them will not show histological nodal invasion. P-glycoprotein expression in the rescue mastectomy specimen has finally been identified as a significant predictor of patient survival. We studied the expression of the genes encoding multidrug resistance associated protein (MDR1) and lung cancer associated resistance protein (LRP) in formalin-fixed, paraffin-embedded tumor samples from 52 patients treated for locally advanced breast cancer by means of induction chemotherapy followed by rescue mastectomy. P-glycoprotein expression was assessed by means of immunohistochemistry before treatment in 23 cases, and by means of reverse-transcriptase-mediated polymerase chain reaction (RT-PCR) after treatment in 46 (6 failed). LRP expression was detected by means of immunohistochemistry, with the LRP-56 monoclonal antibody, in 31 cases before treatment. Immunohistochemistry for detecting the expression of c-erb-B2, p53, Ki67, estrogen receptor and progesterone receptor are routinely performed in our laboratory in every case, and the results obtained were included in the study. All patients had received between two and six cycles of standard 5-fluorouracil, doxorubicin and cyclophosphamide (FAC) chemotherapy, with two exceptions [one patient received four cycles of a docetaxel-adriamycin combination, and the other four cycles of standard cyclophosphamide-methotrexate-5-fluorouracil (CMF) polychemotherapy]. Response was assessed in accordance with the Response Evaluation Criteria In Solid Tumors (RECIST). By these, 2 patients achieved a complete clinical response, 37 a partial response, and the remaining 13 showed stable disease. This makes a total clinical response rate of 75.0%. None achieved a complete pathological response. < 0.05). Finally, in a logistic regression multivariate model, histology other than ductal, a Ki67 labeling index of at least 20% and the combination of LRP and MDR1 positivity emerged as independent predictors of axillary node invasion at the time of rescue mastectomy. The expression of different genes involved in resistance to chemotherapy, both before and after treatment with neoadjuvant, is associated with the presence of axillary node invasion at rescue surgery in locally advanced breast cancer. This might reflect the presence of intrinsically resistant clones before any form of therapy, which persist after it, and could be helpful both for prognosis and for the choice of individual treatment.

Evidence-based medicine (EBM) is being embraced by an increasing number of practitioners and advocates of complementary and alternative medicine (CAM). A significant constituency within CAM, however, appears to have substantive doubts about EBM and some are expressly hostile. Many of the arguments raised against EBM within the CAM community are based on a caricature radically at odds with established, accepted and published principles of EBM practice. Contrary to what has sometimes been argued, EBM is not cookbook medicine that ignores individual needs. Neither does EBM mandate that only proven therapies should be used. Before EBM, decisions on health care tended to be based on tradition, power and influence. Such modes usually act to the disadvantage of marginal groups. By placing CAM on an equal footing with conventional medicine - what matters for both is evidence of effectiveness - EBM provides an opportunity for CAM to find an appropriate and just place in health care.

NFATp is one member of a family of transcriptional activators whose nuclear accumulation and hence transcriptional activity is regulated in mammalian cells. Human NFATp exists as a phosphoprotein in the cytoplasm of naive T cells. Upon antigen stimulation, NFATp is dephosphorylated, accumulates in nuclei, and functions to regulate transcription of genes including those encoding cytokines. While the properties of the DNA binding domain of NFATp have been investigated in detail, biochemical studies of the transcriptional activation and regulated association with nuclei have remained unexplored because of a lack of full length, purified recombinant NFATp. We developed methods for expressing and purifying full length recombinant human NFATp that has all of the properties known to be associated with native NFATp. The recombinant NFATp binds DNA on its own and cooperatively with AP-1 proteins, activates transcription in vitro, is phosphorylated, can be dephosphorylated by calcineurin, and exhibits regulated association with nuclei in vitro. Importantly, activation by recombinant NFATp in a reconstituted transcription system required regions of the protein outside of the central DNA binding domain. We conclude that NFATp is a bona fide transcriptional activator. Moreover, the reagents and methods that we developed will facilitate future studies on the mechanisms of transcriptional activation and nuclear accumulation by NFATp, a member of an important family of transcriptional regulatory proteins.

was diagnosed by mammography in 78.9% and by MR mammography in 68.4% of patients. Combination of all three diagnostic methods lead to the best results for detection of invasive cancer and multifocal disease. However, sensitivity of mammography and sonography combined was identical to that of MR mammography (ie 94.6%).

. Despite demonstrable chemopreventive efficacy in this model, none of these agents, alone or in combination, had any significant impact on the expression of TGF-βs in the mammary ductal epithelium or periductal stroma as determined by immunohistochemistry. These data suggest that TGF-βs are not likely to be useful biomarkers of chemopreventive efficacy in a clinical setting.

To describe the design of the Million Women Study and the characteristics of the study population. Population-based cohort study of women aged 50-64 in the UK. Women are asked to join the Million Women Study when they are invited to routine screening for breast cancer at 61 of the screening centres of the UK National Health Service Breast Screening Programme (NHSBSP). An estimated 71% of women screened by the NHSBSP return a completed questionnaire. 800 000 women were recruited between May 1996 and June 1999, and it is planned that an additional 200 000 will be recruited by the year 2000. The characteristics of the first 121 000 women recruited into the Million Women Study are described here. At recruitment 33% of the study population were currently using hormone replacement therapy and 47% had used it at some time. Over half (54%) had used oral contraceptives, and 18% were current smokers at the time of recruitment. Before they were screened 1.4% of the women had been diagnosed with breast cancer in the past, 6% had a mother with a history of breast cancer and 3.7% had a sister with a history of breast cancer. It is estimated that 1 million women will have been recruited by early in the year 2000, and that by the end of the year 2002 there will be 5000 screen-detected breast cancers and 23 000 deaths in the cohort, the majority of which will be attributed to cancer (12 600 deaths) and circulatory disease (8000 deaths). By the end of the year 2002, the Million Women Study will have sufficient statistical power to detect relative risks of 0.8 or less, or of 1.2 or more in current users compared with never users of hormone replacement therapy for mortality from breast cancer, colorectal cancer, lung and ovarian cancer, ischaemic heart disease and stroke.

gene in 50-60% of individuals. Several studies have demonstrated a possible link with the GSTM1-null genotype and susceptibility to cancer. Furthermore, a GSTM1 isoenzyme has been positively associated with protective effect against mutagenic drugs, such as alkylating agents and anthracyclines. To determine whether GSTM1 polymorphisms are associated with tumour characteristics and survival in advanced breast cancer patients, and whether it may constitute a prognostic factor. We genotyped 92 patients receiving primary chemotherapy, which included cyclophosphamide, doxorubicine and 5-fluorouracil. The relationships between allelism at GSTM1 and clinicopathological parameters including age, menopausal status, tumour size, grade hormone receptors, involved nodes and p53 gene mutations were analysed. Of the patients with GSTM1-positive genotype, tissue samples obtained before and after treatment were available from 28 cases, allowing RNA extraction and GSTM1 expression by reverse transcription polymerase chain reaction. Relationships with clinical response to chemotherapy, and disease-free and overall survival were also evaluated. The data obtained was analysed using logistic regression to estimate the odds ratio and 95% confidence interval. = 0.0086) were the only factors associated with reduced disease-free or overall survival. GSTM1-null genotype alone had no effect on tumour characteristics and outcome of patients with advanced breast cancers. The lack of correlation of GSTM1 genotype with clinical tumour features, clinical response to chemotherapy and survival exclude a role for GSTM1 polymorphism as a prognostic factor in advanced breast cancer.

Resistance to mammary tumorigenesis in Copenhagen rats is associated with loss of early preneoplastic lesions known as intraductal proliferations. The cause of this disappearance, however, is unknown. protein expression, bromodeoxyuridine labeling and apoptotic indices, or mast cell infiltration between Copenhagen and Wistar-Furth lesions at any time. expression in Copenhagen intraductal proliferations may play a role in the resistance of these rats to mammary tumorigenesis.

expressing dousble-stranded RNA corresponding to a specific gene. An optimized feeding method is presented that results in phenotypes at least as strong as those produced by direct injection of RNA for embryonic lethal genes, and stronger for genes with post-embryonic phenotypes. expressing dsRNA corresponding to a specific gene; this mode of dsRNA introduction is conventionally considered to be less efficient than direct injection, however, and has therefore seen limited use, even though it is considerably less labor-intensive. and a gene encoding a MADS-box protein) and one gene encoding a novel protein that results in a high-incidence-of-males phenotype. RNAi by feeding can provide significant information about the functions of an individual gene beyond that provided by injection. Moreover, it can be used for special applications for which injection or the use of mutants is sometimes impracticable (for example, titration, biochemistry and large-scale screening). Thus, RNAi by feeding should make possible new experimental approaches for the use of genomic sequence information.

A new humidifier for use during mechanical ventilation in endotracheally intubated patients is described and tested. The humidifier is based on a heat-moisture exchanger, which absorbs the expired heat and moisture and releases it into the inspired air. External heat and water are then added at the patient side of the heat-moisture exchanger, so that the inspired gas should reach 100% humidity (44 mg/l) at 37°C. In bench tests using constant and decelerating inspiratory flow and minute volumes of 3–25 l the device gave an absolute humidity of 41–44 mg/l, and it reduced the amount of water consumed in eight mechanically ventilated patients compared with a conventional active humidifier. During a 24-h test period there was no water condensation in the ventilator tubing with the new device. Devices for active humidification of the inspired air in mechanically ventilated patients cause water condensation in the ventilator tubing, which may become contaminated or interfere with the function of the ventilator. The present study describes and tests the performance of a new humidifier, which is designed to eliminate water condensation. To test the performance of the new humidifier at different ventilator settings in a lung model, and to compare this new humidifier with a conventional active humidifier in ventilator-treated critically ill patients. O/l air). The external water is delivered to the humidification device via a pump onto a wick and then evaporated into the inspired air by an electrical heater. The microprocessor controls the water pump and the heater by an algorithm using the minute ventilation (which is fed into the microprocessor) and the airway temperature measured by a sensor mounted in the flex-tube on the patient side of the humidification device. The performance characteristics were tested in a lung model ventilated with a constant flow (inspiratory:expiratory ratio 1:2, rate 12–20 breaths/min and a minute ventilation of 3–25 l/min) or with a decelerating flow (inspiratory:expiratory ratio 1:2, rate 12–15 breaths/min and a minute ventilation of 4.7–16.4 l/min). The device was also tested prospectively and in a randomized order compared with a conventional active humidifier (Fisher & Paykel MR730, Auckland, New Zealand) in eight mechanically ventilated, endotracheally intubated patients in the intensive care unit. The test period with each device was 24 h. The amount of fluid consumed and the amount of water in the water traps were measured. The number of times that the water traps were emptied, changes of machine filters, the suctions and quality of secretions, nebulizations, and the amount of saline instillations and endotracheal tube obstruction were recorded. In order to evaluate increased expiratory resistance due to the device, the airway pressure was measured at the end of a prolonged end-expiratory pause at 1 h of use and at the end of the test, and was compared with the corresponding pressure before the experiment. The body temperature of the patient was measured before and after the test of each device. < 0.0008). The same relations were found when the water consumption was corrected for differences in minute ventilation. The new humidifier, the Humid-Heat, gave an absolute humidity of 41–44 mg/l at 37°C in the bench tests. The tests in ventilated patients showed that the device was well tolerated and that condensation in the tubing was eliminated. There was no need to empty water traps. The test period was too short to evaluate whether the new device had any other advantages or disadvantages compared with conventional humidifiers.

Current smoking was strongly and inversely associated with high-risk patterns, after adjustment for concomitant risk factors. Relative to never smokers, current smokers were significantly less likely to have a high-risk pattern. Similar results were obtained when the analysis was confined to postmenopausal women. Past smoking was not related to the mammographic parenchymal patterns. The overall effect in postmenopausal women lost its significance when adjusted for other risk factors for P2/DY patterns that were found to be significant in the present study, although the results are still strongly suggestive. The present data indicate that adjustment for current smoking status is important when evaluating the relationship between mammographic parenchymal pattern and breast cancer risk. They also indicate that smoking is a prominent potential confounder when analyzing effects of other risk factors such as obesity-related variables. It appears that parenchymal patterns may act as an informative biomarker of the effect of cigarette smoking on breast cancer risk. ]. There are no published studies that assessed the relationship between smoking and mammographic parenchymal patterns. ], the association between smoking habits and mammographic parenchymal patterns are examined. The full results will be published elsewhere. Study subjects were members of the EPIC cohort in Norwich who also attended the prevalence screening round at the Norwich Breast Screening Centre between November 1989 and December 1997, and were free of breast cancer at that screening. Cases were defined as women with a P2/DY Wolfe's mammographic parenchymal pattern on the prevalence screen mammograms. A total of 203 women with P2/DY patterns were identified as cases and were individually matched by date of birth (within 1 year) and date of prevalence screening (within 3 months) with 203 women with N1/P1 patterns who served as control individuals. Two views, the mediolateral and craniocaudal mammograms, of both breasts were independently reviewed by two of the authors (ES and RW) to determine the Wolfe mammographic parenchymal pattern. ]. In the present study we examined the subjects' personal history of benign breast diseases, menstrual and reproductive factors, oral contraception and hormone replacement therapy, smoking, and anthropometric information such as body mass index and waist:hip ratio. ], and were adjusted for possible confounding factors. . Cases were leaner than controls. A larger percentage of cases were nulliparous, premenopausal, current hormone replacement therapy users, had a personal history of benign breast diseases, and had had a hysterectomy. A larger proportion of controls had more than three births and were current smokers. shows the unadjusted and adjusted OR estimates for Wolfe's high-risk mammographic parenchymal patterns and smoking in the total study population and in postmenopausal women separately. Current smoking was strongly and inversely associated with high-risk patterns, after adjustment for concomitant risk factors. Relative to never smokers, current smokers were significantly less likely to have a high-risk pattern (OR 0.37, 95% CI 0.14-0.94). Similar results were obtained when the analysis was confined to postmenopausal women. Past smoking was not related to mammographic parenchymal patterns. The overall effect in postmenopausal women lost its significance when adjusted for other risk factors for P2/DY patterns that were found to be significant in the present study, although the results were still strongly suggestive. There was no interaction between cigarette smoking and body mass index. ] found a reduced risk of breast density in association with smoking, although the magnitude of the reduction was unclear. The present findings suggest that this reduction is large. ], who reported that breast cancer risk in younger women (younger than 45 years) may be reduced among current smokers who began smoking at an early age (OR 0.59, 95% CI 0.41-0.85 for age 15 years or younger) and among long-term smokers (OR 0.70, 95% CI 0.52-0.94 for those who had smoked for 21 years or more). ], however, do not support these findings. ]. The present data indicate that adjustment for current smoking status is important when evaluating the relationship between mammographic parenchymal pattern and breast cancer risk. They also indicate smoking as a prominent potential confounder when analyzing effects of other risk factors such as obesity-related variables. It seems that parenchymal patterns may act as an informative biomarker of the effect of cigarette smoking on breast cancer risk.

together with the effects of overexpression of IDO on the growth and morphology of cells. Overexpression of IDO in the murine macrophage cell line RAW 264.7 and the murine fibrosarcoma cell line MC57, resulted in the growth of macroscopic cell foci, with altered cell adhesion properties. The expression of IDO was also detected during adhesion of wild type, nontransfected cells in tissue culture to standard cell growth substrates. Inhibition of this expression, likewise resulted in alterations in cell adhesion. Overexpression of IDO or inhibition of endogenous IDO expression was accompanied by changes in metalloproteinase expression and also in the expression and activity of the cyclooxygenase enzymes. In the case of RAW cells, IDO effects on cell growth could be reversed by adding back prostaglandins. These results suggest that catabolism of the rarest essential amino acid may regulate processes such as cell adhesion and prostaglandin synthesis.

The known association between the acute-phase response and progressive joint damage may be the direct result of synovial A-SAA-induced effects on cartilage degradation. encodes constitutive SAA and is minimally inducible. A-SAA increases dramatically during acute inflammation and may reach levels that are 1000-fold greater than normal. A-SAA is mainly synthesized in the liver, but extrahepatic production has been demonstrated in many species, including humans. A-SAA mRNA is expressed in RA synoviocytes and in monocyte/macrophage cell lines such as THP-1 cells, in endothelial cells and in smooth muscle cells of atherosclerotic lesions. A-SAA has also been localized to a wide range of histologically normal tissues, including breast, stomach, intestine, pancreas, kidney, lung, tonsil, thyroid, pituitary, placenta, skin and brain. To identify the cell types that produce A-SAA mRNA and protein, and their location in RA synovium. Rheumatoid synovial tissue was obtained from eight patients undergoing arthroscopic biopsy and at joint replacement surgery. Total RNA was analyzed by reverse transcription (RT) polymerase chain reaction (PCR) for A-SAA mRNA. PCR products generated were confirmed by Southern blot analysis using human A-SAA cDNA. Localization of A-SAA production was examined by immunohistochemistry using a rabbit antihuman A-SAA polyclonal antibody. PrimaryRA synoviocytes were cultured to examine endogenous A-SAA mRNA expression and protein production. demonstrates RT-PCR products generated using synovial tissue from three representative RA patients. Analysis of RA synovial tissue revealed differences in A-SAA mRNA levels between individual RA patients. ). This study demonstrates that A-SAA mRNA is expressed in several cell populations infiltrating RA synovial tissue. A-SAA mRNA expression was observed in all eight unseparated RA tissue samples studied. A-SAA mRNA expression and protein production was demonstrated in primary cultures of purified RA synoviocytes. Using immunohistochemical techniques, A-SAA protein appeared to colocalize with both lining layer and sublining layer synoviocytes, macrophages and some endothelial cells. The detection of A-SAA protein in culture media supernatants harvested from unstimulated synoviocytes confirms endogenous A-SAA production, and is consistent with A-SAA mRNA expression and translation by the same cells. Moreover, the demonstration of A-SAA protein in RA synovial tissue, RA cultured synoviocytes, macrophages and endothelial cells is consistent with previous studies that demonstrated A-SAA production by a variety of human cell populations. The RA synovial lining layer is composed of activated macrophages and fibroblast-like synoviocytes. The macrophage is the predominant cell type and it has been shown to accumulate preferentially in the surface of the lining layer and in the perivascular areas of the sublining layer. Nevertheless, our observations strongly suggest that A-SAA is produced not only by synoviocytes, but also by synovial tissue macrophage populations. Local A-SAA protein production by vascular endothelial cells was detected in some, but not all, of the tissues examined. The reason for the variability in vascular A-SAA staining is unknown, but may be due to differences in endothelial cell activation, events related to angiogenesis or the intensity of local inflammation. The value of measuring serum A-SAA levels as a reliable surrogate marker of inflammation has been demonstrated for several diseases including RA, juvenile chronic arthritis, psoriatic arthropathy, ankylosing spondylitis, Behçet's disease, reactive arthritis and Crohn's disease. It has been suggested that serum A-SAA levels may represent the most sensitive measurement of the acute-phase reaction. In RA, A-SAA levels provide the strongest correlations with clinical measurements of disease activity, and changes in serum levels best reflect the clinical course. A number of biologic activities have been described for A-SAA, including several that are relevant to the understanding of inflammatory and tissue-degrading mechanisms in human arthritis. A-SAA induces migration, adhesion and tissue infiltration of circulating monocytes and polymorphonuclear leukocytes. In addition, human A-SAA can induce interleukin-1β, interleukin-1 receptor antagonist and soluble type II tumour necrosis factor receptor production by a monocyte cell line. Moreover, A-SAA can stimulate the production of cartilage-degrading proteases by both human and rabbit synoviocytes. The effects of A-SAA on protease production are interesting, because in RA a sustained acute-phase reaction has been strongly associated with progressive joint damage. The known association between the acute-phase response and progressive joint damage may be the direct result of synovial A-SAA-induced effects on cartilage degradation. In contrast to noninflamed synovium, A-SAA mRNA expression was identified in all RA tissues examined. A-SAA appeared to be produced by synovial tissue synoviocytes, macrophages and endothelial cells. The observation of A-SAA mRNA expression in cultured RA synoviocytes and human RA synovial tissue confirms and extends recently published findings that demonstrated A-SAA mRNA expression in stimulated RA synoviocytes, but not in unstimulated RA synoviocytes.

The iridocorneal angle forms in the mammalian eye from undifferentiated mesenchyme between the root of the iris and cornea. A major component is the trabecular meshwork, consisting of extracellular matrix organized into a network of beams, covered in trabecular endothelial cells. Between the beams, channels lead to Schlemm's canal for the drainage of aqueous humor from the eye into the blood stream. Abnormal development of the iridocorneal angle that interferes with ocular fluid drainage can lead to glaucoma in humans. Little is known about the precise mechanisms underlying angle development. There are two main hypotheses. The first proposes that morphogenesis involves mainly cell differentiation, matrix deposition and assembly of the originally continuous mesenchymal mass into beams, channels and Schlemm's canal. The second, based primarily on rat studies, proposes that cell death and macrophages play an important role in forming channels and beams. Mice provide a potentially useful model to understand the origin and development of angle structures and how defective development leads to glaucoma. Few studies have assessed the normal structure and development of the mouse angle. We used light and electron microscopy and a cell death assay to define the sequence of events underlying formation of the angle structures in mice. The mouse angle structures and developmental sequence are similar to those in humans. Cell death was not detectable during the period of trabecular channel and beam formation. These results support morphogenic mechanisms involving organization of cellular and extracellular matrix components without cell death or atrophy.

An A54T polymorphism at the fatty acid binding protein 2 (FABP2) locus was found to be associated with insulin resistance in non-diabetic Pima Indians. To see whether this association is present in other populations, we performed a cross sectional study to examine the role of this polymorphism on insulin resistance in 55 healthy and normotensive Caucasian subjects with normal glucose tolerance. Insulin sensitivity (%S) and beta cell function (%B) were assessed using the Homeostasis Model Assessment (HOMA). Their genotypes were determined using a polymerase chain reaction-restriction fragment length polymorphism assay. The relationship between the genotypes and the phenotypes was examined. After genotyping, we identified 24 AA, 27 AT and 4 TT subjects. The TT subjects were combined with the AT subjects during the analysis due to its small sample size. No differences were noted in gender distribution, clinical features, and fasting lipid profile between the two genotypic groups (AA vs. AT/TT). The AT/TT group had a higher fasting plasma insulin concentration and a lower %S than the AA group (p = 0.0444 and p = 0.0461, respectively). However, no differences were noted in plasma glucose concentrations and %B. Univariate analysis revealed that this polymorphism explained 7.3% of the variation in %S. Multivariate analysis revealed that the polymorphism was an independent determinant for %S (p = 0.0434) and with body mass index accounted for 28.7% of the variation in %S. In contrast, this polymorphism had no impact on %B. The A54T polymorphism at the FABP2 locus is a risk factor for insulin resistance in a Caucasian population.

Destruction of cartilage and bone are hallmarks of human rheumatoid arthritis (RA), and controlling these erosive processes is the most challenging objective in the treatment of RA. Systemic interleukin-4 treatment of established murine collagen-induced arthritis suppressed disease activity and protected against cartilage and bone destruction. Reduced cartilage pathology was confirmed by both decreased serum cartilage oligomeric matrix protein (COMP) and histological examination. In addition, radiological analysis revealed that bone destruction was also partially prevented. Improved suppression of joint swelling was achieved when interleukin-4 treatment was combined with low-dose prednisolone treatment. Interestingly, synergistic reduction of both serum COMP and inflammatory parameters was noted when low-dose interleukin-4 was combined with prednisolone. Systemic treatment with interleukin-4 appeared to be a protective therapy for cartilage and bone in arthritis, and in combination with prednisolone at low dosages may offer an alternative therapy in RA. . Of great importance is that IL-4 could not be detected in synovial fluid or in tissues. This absence of IL-4 in the joint probably contributes to the disturbance in the Th1/Th2 balance in chronic RA. Collagen-induced arthritis (CIA) is a widely used model of arthritis that displays several features of human RA. Recently it was demonstrated that the onset of CIA is under stringent control of IL-4 and IL-10. Furthermore, it was demonstrated that exposure to IL-4 during the immunization stage reduced onset and severity of CIA. However, after cessation of IL-4 treatment disease expression increased to control values. Because it was reported that IL-4 suppresses several proinflammatory cytokines and matrix degrading enzymes and upregulates inhibitors of both cytokines and catabolic enzymes, we investigated the tissue protective effect of systemic IL-4 treatment using established murine CIA as a model. Potential synergy of low dosages of anti-inflammatory glucocorticosteroids and IL-4 was also evaluated. DBA-1J/Bom mice were immunized with bovine type II collagen and boosted at day 21. Mice with established CIA were selected at day 28 after immunization and treated for days with IL-4, prednisolone, or combinations of prednisolone and IL-4. Arthritis score was monitored visually. Joint pathology was evaluated by histology, radiology and serum cartilage oligomeric matrix protein (COMP). In addition, serum levels of IL-1Ra and anticollagen antibodies were determined. ). Systemic IL-4 administration increased serum IL-1Ra levels and reduced anticollagen type II antibody levels. Treatment with low-dose IL-4 (0.1 μg/day) was ineffective in suppressing disease score, serum COMP or joint destruction. Synergistic suppression of both arthritis severity and COMP levels was noted when low-dose IL-4 was combined with prednisolone (0.05 mg/kg/day), however, which in itself was not effective. In the present study, we demonstrate that systemic IL-4 treatment ameliorates disease progression of established CIA. Although clinical disease progression was only arrested and not reversed, clear protection against cartilage and bone destruction was noted. This is in accord with findings in both human RA and animal models of RA that show that inflammation and tissue destruction sometimes are uncoupled processes. Of great importance is that, although inflammation was still present, strong reduction in serum COMP was found after exposure to IL-4. This indicated that serum COMP levels reflected cartilage damage, although a limited contribution of the inflamed synovium cannot be excluded. Increased serum IL-1Ra level (twofold) was found after systemic treatment with IL-4, but it is not likely that this could explain the suppression of CIA. We and others have reported that high dosages of IL-1Ra are needed for marked suppression of CIA. As reported previously, lower dosages of IL-4 did not reduce clinical disease severity of established CIA. Of importance is that combined treatment of low dosages of IL-4 and IL-10 appeared to have more potent anti-inflammatory effects, and markedly protected against cartilage destruction. Improved anti-inflammatory effect was achieved with IL-4/prednisolone treatment. In addition, synergistic effects were found for the reduction of cartilage and bone destruction. This indicates that systemic IL-4/prednisolone treatment may provide a cartilage and bone protective therapy for human RA. < 0.05, versus control, by Mann-Whitney U test.

Although the role of the osteoclast in bone resorption is becoming better understood, much remains to be learned about osteoclastogenesis and the exact mechanism of action of anti-resorbing agents such as 17β-estradiol. This study investigated bone and morphologic osteoclast alterations following long-term estrogen administration to the B6D2F1 mouse. B6D2F1 mice aged 4-5 weeks were exposed to high levels of estrogen via implanted silastic tubing for at least 12 weeks; controls received empty tubing. Femurs of control and treated mice were assessed with radiology, quantitative histomorphometry and transmission electron microscopy. After 8 weeks of treatment, there was radiologic evidence of severe osteosclerosis and 86% of femoral marrow space was replaced with bone. After 12 weeks histologic studies of treated animals revealed that osteoclasts were positive for tartrate-resistant acid phosphatase but showed markedly abnormal ultrastructure which prevented successful bone resorption. Findings extend our understanding of osteoclast structure and function in the mouse exposed in vivo to high doses of estrogen. Ultrastructural examination showed that osteoclasts from estrogen-treated mice were unable to seal against the bone surface and were unable to form ruffled borders.

The regulation of programmed cell death is critical to developmental homeostasis and normal morphogenesis of embryonic tissues. Survivin, a member of the inhibitors of apoptosis protein (IAP) family primarily expressed in embryonic cells, is both an anti-apoptosis and a pro-survival factor. Since our previous studies have demonstrated the importance of apoptosis during embryonic submandibular salivary gland (SMG) development, we postulated that survivin is a likely mediator of SMG epithelial cell survival. Stage SMGs. Survivin is known to be a pro-survival and anti-apoptotic factor. Given that survivin translocation into the nucleus is required for the induction of entry into the cell cycle and the inhibition of apoptosis, our demonstration of nuclear-localized survivin protein in presumptive ductal and proacinar lumen-bounding cells suggests that survivin may be a key mediator of embryonic SMG epithelial cell survival.

gene in type 1 diabetic patients with (n = 82) and without DN (n = 84). gene were significantly lower in DN patients than in diabetic subjects without DN. In contrast, the Val allele (49.4% vs. 31.5%, p < 0.002) and the Val/Val genotype (15.9% vs. 2.4%, p < 0.01) were significantly more frequent in the DN patients than in the control group. gene was associated with DN in a Russian population

] prompted this investigation. ]. sp. WH8113 provides new constraints on its motile mechanism. The spicules are well positioned to transduce energy at the cell membrane into mechanical work at the cell surface. One model is that an unidentified motor embedded in the cell membrane utilizes the spicules as oars to generate a traveling wave external to the surface layer in the manner of ciliated eukaryotes.

The striatum plays a pivotal role in modulating motor activity and higher cognitive function. We analyzed variation in striatal volume and neuron number in mice and initiated a complex trait analysis to discover polymorphic genes that modulate the structure of the basal ganglia. ) and is associated with differences of up to 400,000 neurons among animals. We have discovered remarkable numerical and volumetric variation in the mouse striatum, and we have been able to map two QTLs that modulate independent anatomic parameters.

R3. R3, PLCβ2, and γ13 positive cells. receptor expressed in taste cells and our data suggest it plays an important role in bitter taste transduction.

In Brazil coronary heart disease (CHD) constitutes the most important cause of death in both sexes in all the regions of the country and interestingly, the difference between the sexes in the CHD mortality rates is one of the smallest in the world because of high rates among women. Since a question has been raised about whether or how the incidence of several CHD risk factors differs between the sexes in Brazil the prevalence of various risk factors for CHD such as high blood cholesterol, diabetes mellitus, hypertension, obesity, sedentary lifestyle and cigarette smoking was compared between the sexes in a Brazilian population; also the relationships between blood cholesterol and the other risk factors were evaluated. The population presented high frequencies of all the risk factors evaluated. High blood cholesterol (CHOL) and hypertension were more prevalent among women as compared to men. Hypertension, diabetes and smoking showed equal or higher prevalence in women in pre-menopausal ages as compared to men. Obesity and physical inactivity were equally prevalent in both sexes respectively in the postmenopausal age group and at all ages. CHOL was associated with BMI, sex, age, hypertension and physical inactivity. In this population the high prevalence of the CHD risk factors indicated that there is an urgent need for its control; the higher or equal prevalences of several risk factors in women could in part explain the high rates of mortality from CHD in females as compared to males.

and some other genomes. Here, we apply the comparative genomic approach to the prediction of the ArgR-binding sites in all completely sequenced bacterial genomes. Candidate arginine repressor binding sites were identified upstream of arginine transport and metabolism genes. ) belonging to the candidate arginine regulons.

as a reporter, in that they can be easily visualized without recourse to the vital substrates required to visualize β-gal in living tissue. ) pattern. The resulting EYFP or ECFP expression patterns indicated that the reporter strains function as faithful monitors of Cre activity. expression cannot easily be detected in living tissue, the EYFP and ECFP reporter strains are useful for monitoring the expression of Cre and tracing the lineage of these cells and their descendants in cultured embryos or organs. The non-overlapping emission spectra of EYFP and ECFP make them ideal for double labeling studies in living tissues.

< 0.001). Of the 101 patients who were positive for at least one of these four autoantibodies, 57% were positive for only one. Finally, anti-SA identified a subset of predominantly male RA patients with severe, erosive disease. Anti-SA, AFA and anti-CCP are all specific for early RA but, overall, have little additional diagnostic value over RF alone. Although these antibodies may preferentially recognize citrullinated antigens, the modest degree of concordance between them in individual patient sera suggests that it is unlikely a single antigen is involved in generating these responses. A spectrum of autoantibodies is now known to be specifically associated with RA. There continues to be uncertainty as to what stage of the disease each of these autoantibodies develop, and whether they are associated with unique clinical features. To help address these questions, a spectrum of autoantibodies known to be associated with RA in a cohort of patients with early synovitis was evaluated. An inception cohort of 238 patients having peripheral joint synovitis of less than 12 months duration was evaluated clinicially then followed prospectively for 1 year. Patients were classified as having RA on the basis of fulfilling the 1987 criteria. Serum samples collected at the time of the initial evaluation were tested for anti-Sa and anti-RA-33 using immunoblotting, and to (pro)filaggrin (AFA), anti-CCP, and calpastatin (anti-RA-1) using enzyme-linked immunosorbent assay techniques. AKA were detected using immunoflurescence on human epidermal tissue. RF was tested by nephelometry. HLA-DRB1 alleles were determined using sequence specific primers. Initial and 1 year radiographs were evaluated for the presence of erosions. < 0.001). Despite this high level of correlation, of the 101 patients who were positive for at least one of these four autoantibodies, 57% were positive for only one, suggesting considerable variability in individual reactivity patterns. , the presence of each of these autoantibodies was significantly associated with having two shared epitope alleles, even when only the RA patients were considered. =0.03). Neither RF nor SE were associated with the disease severity measures, and analyses evaluating all the other autoantibodies failed to reveal a similar trend. Despite a well-documented lack of specificity, RF continues to be a central part of the definition of RA, primarily because of its favourable sensitivity profile. In our cohort, RF had a sensitivity of 66%, a specificity of 87%, and an overall accuracy of 78% for the diagnosis of RA. AFA, anti-Sa, anti-CCP were all highly specific for this diagnosis, and when any of them were present in conjunction with RF, the specificity for RA approached 100%. Potentially of more importance to the clinician is the diagnostic value of these antibodies when RF is not detectable. Our data indicate that only 31% of RF- RA patients had any of AKA, AFA, anti-Sa or anti-CCP, and that anti-Sa was the most specific for this diagnosis. This modest level of sensitivity suggests that testing for this spectrum of autoantibodies carries little advantage over RF alone in diagnosing early RA. AFA, AKA, and antiperinuclear factor (APF) have all been proposed to identify a common antigen present in the skin protein (pro)filaggrin. It has continued to be puzzling why a skin antigen would be targeted relatively specifically in a disorder that is primarily articular. A potential explanation for this may relate to the demonstration that citrulline appears to be an essential constituent of the antigenic determinants recognized by AKA, APF, and AFA. The citrulline rich (pro)filaggrin molecule makes an ideal substrate for detecting this reactivity. Moreover, the SA antigen, which, unlike (pro)filaggrin, is detectable in rheumatoid synovium, has recently been shown to also be citrullinated. It is thus possible that AKA, AFA, APE, and anti-Sa all recognize one or more citrullinated antigens. Despite this possibility, the modest degree of concordance between them in individual patient sera suggests that it is unlikely that a single antigen is involved in generating these responses. This study provides evidence suggesting that anti-Sa antibodies appear to be a marker for a subset of early RA patients whose disease may be more severe and erosive. Moreover, it was determined that anti-Sa, AFA, and anti-CCP were all highly associated with SE, particularly two copies. We examined a spectrum of potential RA severity indicators including the number of swollen joints, CRP level, and presence of early radiographic erosions. Our data indicate that anti-Sa was more highly associated with these measures of RA severity than any other parameter, including the most accepted prognostic indicators, RF and SE. In conclusion, it is demonstrated that antibodies directed against putatively citrullinated antigens including SA, filaggrin, keratin, and CCP are the most specific for RA, and are detectable early in the disease course. It will be of interest to find out whether the cumulative prevalence of specific autoantibody subsets tends to increase over time, as this would suggest that the mechanisms underlying the development of these reactivities continue to evolve over the course of the arthropathy.

There are numerous prehosital descriptive scoring systems, and it is uncertain whether they are efficient in assessing of the severity of illness and whether they have a prognostic role in the estimation of the illness outcome (in comparison with that of the prognostic scoring system Acute Physiology and Chronic Health Evaluation [APACHE] II). The purpose of the present study was to assess the value of the various scoring systems in predicting outcome in nontraumatic coma patients and to evaluate the importance of mental status measurement in relation to outcome. < 0.05 was considered statistically significant. For prediction of mortality, the best cutoff points were 19 for APACHE II, 18 for MEES and 5 for GCS. The best cutoffs for the Youden index were 0.63 for APACHE II, 0.61 for MEES and 0.65 for GCS. The correct prediction of outcome was achieved in 79.9% for APACHE II, 78.3% for MEES and 81.9% for GCS. The area under the ROC curve (mean ± standard error) was 0.86 ± 0.02 for APACHE II, 0.84 ± 0.06 for MEES and 0.88 ± 0.03 for GCS. There were no statistically significant differences among APACHE II, MEES and GCS scores in terms of correct prediction of outcome, Youden index or area under ROC curve. APACHE II is not much better than prehospital descriptive scoring systems (MEES and GCS). APACHE II and MEES should not replace GCS in assessment of illness severity or in prediction of mortality in nontraumatic coma. For the assessment of mortality, the GCS score provides the best indicator for these patients (simplicity, less time-consuming and effective in an emergency situation.

) were estimated from the oral glucose tolerance test. Beta cell function was estimated using %B from the Homeostasis Model Assessment and insulingenic index (dI/dG). (p=0.089) and %B (p=0.083). Furthermore, it had no independent effect on dI/dG (p=0.135). These data suggest that the G-to-A polymorphism in the hepatic GCK promoter is associated with hepatic insulin resistance in Asian Indians.

An association between mumps-measles-rubella (MMR) vaccination and the onset of symptoms typical of autism has recently been suggested. This has led to considerable concern about the safety of the vaccine. A matched case-control study using data derived form the United Kingdom General Practice Research Database. Children with a possible diagnosis of autism will be identified from their electronic health records. All diagnoses will be validated by a detailed review of hospital letters and by using information derived from a parental questionnaire. Ten controls per case will be selected from the database. Conditional logistic regression will be used to assess the association between MMR vaccination and autism. In addition case series analyses will be undertaken to estimate the relative incidence of onset of autism in defined time intervals after vaccination. The study is funded by the United Kingdom Medical Research Council. Electronic health databases offer tremendous opportunities for evaluating the adverse effects of vaccines. However there is much scope for bias and confounding. The rigorous validation of all diagnoses and the collection of additional information by parental questionnaire in this study are essential to minimise the possibility of misleading results.

mutations in mice lead to the development of a variety of tumors, but mammary carcinomas are infrequently found, especially in mice under the age of six months. function have a growth advantage over cells retaining a wild type allele. Tumors with LOH contained high levels of activated AKT/PKB, a downstream target of the PTEN/PI3K pathway. to induce ductal carcinoma in the mammary gland. This animal model may be useful for testing therapies specific for tumors deregulated in the PTEN/PI3K/AKT pathway.

The most appropriate inotropic agent for use in the newborn is uncertain. Dopamine and epinephrine are commonly used, but have unknown effects during hypoxia and pulmonary hypertension; the effects on the splanchnic circulation, in particular, are unclear. respectively were infused for 1 h at each dose. Results were compared with the 1 h hypoxia values by two-way analysis of variance. ; dopamine had no effect on hepatic arterial flow at any dose. Despite these hemodynamic changes, there were no differences in systemic or splanchnic oxygen extraction or consumption at any dose of dopamine or epinephrine. Epinephrine is more effective than dopamine at increasing cardiac output during hypoxia in this model. Although epinephrine preserves the SAP/PAP ratio, dopamine shows preferential pulmonary vasoconstriction, which might be detrimental if it also occurs during the management of infants with persistent fetal circulation. Dopamine, but not epinephrine, increases portal flow and total hepatic flow during hypoxia.

Macrophages that accumulate in the synovium of rheumatoid arthritis patients play an important role in the pathogenesis of this inflammatory disease. However, the mechanism by which macrophages are attracted into the inflamed synovium and accumulate there has not been completely delineated. The results of this study show that rheumatoid arthritis synovial stromal cells produce the chemokines monocyte chemotactic protein-1 and IL-8, and these have the capacity to attract peripheral monocytes. These results suggest that one of the mechanisms by which macrophages accumulate in the inflamed synovium is by responding to the chemokines produced locally.

We have developed and tested a method for printing protein microarrays and using these microarrays in a comparative fluorescence assay to measure the abundance of many specific proteins in complex solutions. A robotic device was used to print hundreds of specific antibody or antigen solutions in an array on the surface of derivatized microscope slides. Two complex protein samples, one serving as a standard for comparative quantitation, the other representing an experimental sample in which the protein quantities were to be measured, were labeled by covalent attachment of spectrally resolvable fluorescent dyes. , sensitivities sufficient for measurement of many clinically important proteins in patient blood samples. These results suggest that protein microarrays can provide a practical means to characterize patterns of variation in hundreds of thousands of different proteins in clinical or research applications.

. This provides a simple model system for studying activation and transdifferentiation of these cells. The introduction of exogenous DNA into these cells is discussed controversially mainly due to the lack of systematic analysis. Therefore, we examined comparatively five nonviral, lipid-mediated gene transfer methods and adenoviral based infection, as potential tools for efficient delivery of DNA to rat hepatic stellate cells and their transdifferentiated counterpart, i.e. myofibroblasts. Transfection conditions were determined using enhanced green fluorescent protein as a reporter expressed under the transcriptional control of the human cytomegalovirus immediate early gene 1 promoter/enhancer. With the use of chemically enhanced transfection methods, the highest relative efficiency was obtained with FuGENE™6 gene mediated DNA transfer. Quantitative evaluation of representative transfection experiments by flow cytometry revealed that approximately 6% of the rat hepatic stellate cells were transfected. None of the transfection methods tested was able to mediate gene delivery to rat myofibroblasts. To analyze if rat hepatic stellate cells and myofibroblasts are susceptible to adenoviral infection, we have inserted the transgenic expression cassette into a recombinant adenoviral type 5 genome as replacement for the E1 region. Viral particles of this replication-deficient Ad5-based reporter are able to infect 100% of rat hepatic stellate cells and myofibroblasts, respectively. Our results indicate that FuGENE™6-based methods may be optimized sufficiently to offer a feasible approach for gene transfer into rat hepatic stellate cells. The data further demonstrate that adenoviral mediated transfer is a promising approach for gene delivery to these hepatic cells.