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R Menon

Publications and source records attributed to R Menon.

At least 37 records · Page 2Linked to original sources
PubMed1999

fMRI evidence for an inverted face representation in human somatosensory cortex.

We provide evidence that the face component of the somatosensory homunculus is actually upside down rather than right-side up along the central sulcus of the human brain. We pneumatically stimulated the forehead or chin of neurologically intact humans while acquiring fMRI images of somatosensory cortex. During forehead stimulation cortical regions along relatively inferior portions of the postcentral gyrus were most active whereas during chin stimulation relatively superior regions were most active. These data are consistent with an inverted face representation along the central sulcus of the human brain.

Adult↗
PubMed1999

T cell responses to a human cartilage autoantigen in the context of rheumatoid arthritis-associated and nonassociated HLA-DR4 alleles.

OBJECTIVE: To analyze the CD4+ T cell responses to the human cartilage antigen glycoprotein-39 (HCgp-39) in the context of rheumatoid arthritis (RA)-associated (DRalphabeta1*0401) and nonassociated (DRalphabeta1*0402) HLA class II molecules. METHODS: Large numbers of HCgp-39-specific T cell hybridomas were generated following immunization of HLA-DR4/human CD4 transgenic, murine major histocompatibility complex class II deficient mice with native HCgp-39. Fine epitope mapping of DRalphabeta1*0401-and DRalphabeta1*0402-restricted T cell hybridomas was performed using overlapping synthetic peptides. Antigen-specific cytokine production by lymph node T cells was evaluated after immunization with native antigen. Proliferative T cell responses of healthy human subjects were compared with the T cell responses of patients with active RA using HCgp-39 epitopes defined in HLA-DR4 transgenic mice. RESULTS: CD4+ T cells from DRalphabeta1*0401 and DRalphabeta1*0402 transgenic mice identified completely different immunodominant peptide epitopes of HCgp-39, and this was not explained by known DR4-binding motifs or direct peptide-binding studies. DRalphabeta1*0401-restricted, antigen-specific T cells produced significantly more interferon-gamma and tumor necrosis factor a in response to HCgp-39 than did T cells from DRalphabeta1*0402 transgenic mice. Finally, HCgp-39 peptides defined in DRalphabeta1*0401 transgenic mice stimulated T cells from HLA-DR4 positive human subjects and RA patients, but not T cells from HLA-DR4 negative individuals. CONCLUSION: T cell epitopes of HCgp-39 that were defined in HLA-DR4 transgenic mice stimulated T cells from human subjects carrying RA-associated HLA-DR4 alleles. HLA-DR4 molecules may influence the disease process in RA both by presentation of selected peptide epitopes and by promoting the production of proinflammatory cytokines in synovial joints.

Adipokines↗
PubMed1999

Stromelysins in placental membranes and amniotic fluid with premature rupture of membranes.

OBJECTIVE: To determine the expression and site of production of stromelysins in fetal membranes and to measure stromelysin 1 levels in amniotic fluid and amniochorion culture media. METHODS: Amniochorionic membranes were cultured from organ explant. Membranes were stimulated with lipopolysaccharide for 24 hours after a 48-hour preincubation period. Membranes were also collected from women after vaginal deliveries. RNA samples from those tissues were subjected to reverse transcriptase-polymerase chain reaction using primers specific for stromelysin 1, stromelysin 2, stromelysin 3, and matrilysin. In situ hybridization and immunohistochemistry were used to localize stromelysin mRNA and peptide. Levels of stromelysin 1 in culture media and amniotic fluid collected from women with preterm premature rupture of membranes (PROM) and at term with intact membranes were compared using enzyme-linked immunosorbant assay. RESULTS: Amniochorion in culture and from laboring and nonlaboring women expressed all three stromelysins. In situ hybridization showed stromelysin mRNA in amnion, chorion, and extracellular matrix. Immunohistochemical analysis localized stromelysin 1 protein to those same regions. Amniotic fluid levels of stromelysin 1 were higher in preterm PROM amniotic fluids (median 3.2 ng/mL) compared with term deliveries with intact membranes (median 1.3 ng/mL) (P = .02). Lipopolysaccharide stimulation in culture increased the release of stromelysin 1 from fetal membranes compared with control (median 70.35 versus 15.8 ng/mL, respectively, P = .05). CONCLUSION: Human fetal membranes are a source of stromelysins 1, 2, and 3. Increased stromelysin 1 during preterm PROM and in vitro after lipopolysaccharide stimulation suggests a possible effect of that matrix metalloproteinase in PROM.

Amniotic Fluid↗
PubMed1999

MMP/TIMP imbalance in amniotic fluid during PROM: an indirect support for endogenous pathway to membrane rupture.

OBJECTIVE: We theorize that excessive degradation of the fetal membrane extracellular matrix (ECM) by specific matrix metalloproteinases (MMPs) results in preterm premature rupture of the membranes (PROM). Active, inhibitor free MMP2 and 9 (gelatinase A and B respectively) can degrade the amniochorion basement membrane Type IV collagen to initiate rupture. This study examines the levels of the gelatinases and their natural inhibitors (tissue inhibitor of matrix metalloproteinases-TIMPs) in the amniotic fluid during PROM, preterm labor (PTL) and at term. METHODS: A total of 51 AF samples were collected from the following groups of patients. Group 1: Women with PTL and no ROM (n = 16) Group 2: Women with PROM (n = 16) irrespective of labor status Group 3: Women at term with intact membranes undergoing cesarean delivery irrespective of labor status (n = 19). ELISA was used to assay MMP2, MMP9, TIMP1 and TIMP2 levels in the amniotic fluid. The active, TIMP free levels of MMP2 were quantitated by zymography followed by computerized densitometry. Active MMP9 was measured using a bioassay that specifically detects MMP9 activity. Statistical analysis was performed by Tukey-Kramer multiple comparison method. RESULTS: PROM is associated with increased MMP2 levels (mean 2125 ng/ml;) when compared with term (mean 1455 ng/ml; p < 0.01) or PTL where a non significant increase was seen (mean 1862 ng/ml; p = ns). MMP9 levels were higher in PROM (mean 15.03 ng/ml) than at term (mean 1.14 ng/ml; p < 0.001) or PTL (mean 3.75 ng/ml; p < 0.01). TIMP1 levels were slightly increased during PROM (mean 3143 ng/ml) compared to term (mean 1892 ng/ml; p < 0.05) pr PTL where a non significant change was seen (mean 2406 ng/ml; p = ns). TIMP2 levels were decreased in PROM (mean 98 ng/ml) compared with term (mean 176 ng/ml; p < 0.05) and PTL (mean 236 ng/ml; p < 0.001). Active, TIMP free MMP2 levels were increased during PROM (mean 233 pg/ml) compared to those at term (mean 132 pg/ml; p < 0.05) or PTL (mean 132 pg/ml; p < 0.05). Active forms of MMP9 were seen only during PROM (mean 632 pg/ml). CONCLUSION: Active, TIMP free forms of MMP2 and 9 are increased in the amniotic fluid of women with PROM. These MMPs can degrade the amniochorion basement membranes and other ECM components resulting in PROM.

Amniotic Fluid↗
PubMed1998

Relaxation of imprinting in Prader-Willi syndrome.

We describe two Prader-Willi syndrome (PWS) patients who exhibit maternal uniparental disomy (UPD) of chromosome 15 and unusual patterns of gene expression and DNA replication. Both were diagnosed during infancy as having PWS; however, their growth and development were atypical compared with others with this condition. Weight was below normal in the first patient, and height and development were within normal limits in the second individual. Hyperphagia and polyphagia were not evident in either patient. Genotypes at multiple genomic loci, allele-specific methylation, gene expression, and DNA replication were analyzed at D15S9 [ZNF127], D15S63 [PW71], SNRPN, PAR5, IPW, and D15S10 in these patients. The maternal imprint (based on the absence of gene expression, synchronous replication, and methylation of both alleles) was retained at SNRPN in these patients, as is the case in others with UPD. By contrast, cells from the first individual expressed PAR5 and ZNF127, whereas the second expressed a single IPW allele. Asynchronous DNA replication was observed in both patients at all loci, except SNRPN. These findings show that a subset of imprinted genes can be transcribed in some PWS patients with maternal UPD and that asynchronous DNA replication is coordinated with this pattern of gene expression. Relaxed imprinting in these patients is consistent with their milder phenotype.

Adolescent↗
PubMed1998

The effect of transforming growth factor and interleukin-10 on interleukin-8 release by human amniochorion may regulate histologic chorioamnionitis.

OBJECTIVE: Amniochorion is a source of interleukin-8 during infection and inflammation. In this study we investigate the role of 2 immunoinhibitory cytokines, transforming growth factor and interleukin-10, in regulating interleukin-8 production from human fetal membranes and define their mechanism of regulation. STUDY DESIGN: Amniochorion was placed in an organ explant system for 72 hours. Tissues were stimulated with lipopolysaccharide (50 ng/mL), lipopolysaccharide plus transforming growth factor-beta (50/50, 50/100), transforming growth factor-beta (50 and 100 ng/mL), lipopolysaccharide plus interleukin-10 (50/50 and 50/100), and interleukin-10 (50 and 100 ng/mL) in culture. Tissue and media samples were frozen until quantitation of interleukin-8 messenger ribonucleic acid and protein. Quantitation of messenger ribonucleic acid was performed by quantitative competitive polymerase chain reaction and protein by enzyme-linked immunoassay, respectively. RESULTS: Lipopolysaccharide-stimulated tissues produced approximately 6 x 10(6) molecules per microliter of interleukin-8 messenger ribonucleic acid compared with 6 x 10(3) molecules per microliter in controls. Transforming growth factor-beta alone and lipopolysaccharide plus transforming growth factor-beta stimulation produced 6 x 10(5) and 6 x 10(4) molecules of interleukin-8 messenger ribonucleic acid per microliter, respectively. Tissues stimulated with lipopolysaccharide plus 50 ng/mL interleukin-10 produced approximately 600 molecules per microliter of interleukin-8 messenger ribonucleic acid, whereas no amplifiable messenger ribonucleic acid was detected in tissues treated with lipopolysaccharide plus 100 ng/mL interleukin-10. Tissues treated with interleukin-10 alone produced 6 x 10(3) molecules of messenger ribonucleic acid, similar to control levels. Enzyme-linked immunosorbent assay data showed similar levels of interleukin-8 peptide release from lipopolysaccharide and lipopolysaccharide plus transforming growth factor-beta-treated fetal membranes. A dose-dependent decrease in interleukin-8 peptide release was seen in tissues treated with lipopolysaccharide plus interleukin-10, whereas stimulation with transforming growth factor or interleukin-10 alone resulted in interleukin-8 peptide release similar to that of control levels. CONCLUSION: Transforming growth factor-beta seems to have no effect on interleukin-8 protein production in the presence of an infectious agent; however, a drop in messenger ribonucleic acid levels was observed. Interleukin-10 in the presence of lipopolysaccharide showed down-regulation of interleukin-8 messenger ribonucleic acid expression and peptide production. These data suggest that fetal membrane interleukin-8 production can be controlled by interleukin-10 during an infectious process.

Amnion↗
PubMed1998

A role for matrix metalloproteinase-9 in spontaneous rupture of the fetal membranes.

OBJECTIVES: Preterm premature rupture of fetal membranes is responsible for 30% to 40% of preterm deliveries. Fetal membranes are composed primarily of collagen. Matrix metalloproteinases are enzymes capable of degrading extracellular matrix macromolecules, including collagens. Expression of matrix metalloproteinase-9 (gelatinase B, 92 kd) and its tissue inhibitor (tissue inhibitor of metalloproteinase-1) has been localized in amnion and chorion. The objective of this study was to determine whether rupture of fetal membranes and intrauterine infection are associated with changes in the expression of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1. STUDY DESIGN: Two hundred one women in the following categories had amniotic fluid retrieved: (1) preterm labor and intact membranes in the presence (n = 42) or absence (n = 21) of microbial invasion of the amniotic cavity, (2) preterm premature rupture of the membranes with (n = 29) or without (n = 23) microbial invasion of the amniotic cavity, and (3) term gestation with intact membranes (n = 50) or with premature rupture of the membranes (n = 40). Women in groups 1 and 2 were matched for gestational age at amniocentesis. Microbial invasion of the amniotic cavity was defined by a positive amniotic fluid culture for micro-organisms. Matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 were measured with use of sensitive and specific immunoassays that were validated for amniotic fluid. RESULTS: Spontaneous rupture of membranes at term is associated with a significant increase in the amniotic fluid concentrations of matrix metalloproteinase-9 (premature rupture of membranes, no labor: median 3.9 ng/mL, range 2. 7 to 11.1 ng/mL vs no premature rupture of membranes, no labor: median <0.4 ng/mL, range <0.4 to 22.4 ng/mL; P <.001). Patients with preterm premature rupture of the membranes had higher median matrix metalloproteinase-9 concentrations than those with preterm labor and intact membranes who were delivered at term (7.6 ng/mL, range <0.4 to 230.81 ng/mL vs <0.4 ng/mL, range <0.4 to 1650 ng/mL; P =.06). Women with microbial invasion of the amniotic cavity had higher median matrix metalloproteinase-9 concentrations than did those without microbial invasion regardless of membrane status (preterm labor: 54.5 ng/mL, range <0.4 to 3910 ng/mL vs <0.4 ng/mL, range <0. 4 to 1650 ng/mL; P <.01; preterm premature rupture of membranes: 179. 8 ng/mL, range <0.4 to 611 ng/mL vs 7.6 ng/mL, range <0.4 to 230.81; P <.001). CONCLUSION: Our data support a role for matrix metalloproteinase-9 in the mechanisms responsible for membrane rupture in term and preterm gestations.

Amnion↗
PubMed1998

IL-15, a novel cytokine produced by human fetal membranes, is elevated in preterm labor.

PROBLEM: Interleukin (IL)-15 is a novel cytokine known to have functions similar to those of IL-2 in the cell-mediated immune response. The objectives of this study were to determine whether IL-15 levels change in labor or preterm labor and to identify the regulatory agents and the site of production of IL-15. METHOD OF STUDY: Amniochorionic membranes were cultured in an organ explant system and were stimulated with lipopolysaccharides (LPSs). Samples were subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) using specific primers for IL-15 and IL-2. The localization of mRNA and protein was accomplished by in situ hybridization and immunocytochemistry. IL-15 was measured in culture media and amniotic fluid from term and preterm gestations by enzyme-linked immunoadsorbent assay (ELISA). RESULTS: RT-PCR indicated the expression of IL-15 mRNA in the amniochorion. In situ hybridization and immunocytochemistry documented that mRNA and peptide for IL-15 are found in amnion, chorion, and decidual cells. ELISA results indicated no significant increase of IL-15 peptides in the culture media after LPS stimulation. Maximum levels of this cytokine were seen in the amniotic fluid (AF) of women with preterm labor compared to term labor. AF levels were not higher in preterm-labor patients with proved infection compared with those without infection. RT-PCR-based detection also showed the presence of two isoforms of IL-15 mRNA known to code for two different leader peptide sequences. IL-2 mRNA expression was not observed in the fetal membranes. CONCLUSIONS: The presence of IL-15 mRNA and peptide in the amniochorion and decidua and its increased presence in the AF during preterm labor suggests a possible role for IL-15 in preterm labor. Amniochorion is also shown to possess two IL-15 isoform leader sequences, the differential expression of which may be involved in the regulation of IL-15 secretion.

Amnion↗
PubMed1998

Presence of four tissue inhibitors of matrix metalloproteinases (TIMP-1, -2, -3 and -4) in human fetal membranes.

PROBLEM: Matrix metalloproteinases play a critical role in fetal membrane extracellular matrix (ECM) homeostasis. Remodeling of the ECM during normal placental development is a balanced activity between various matrix metalloproteinases and their tissue-specific counter-regulatory proteins (tissue inhibitors of matrix metalloproteinases [TIMPs]). We have reported the presence of TIMP-1 and TIMP-2 in placental membranes in culture. In this study we have investigated the membrane expression of TIMP-1 and TIMP-2 during labor and nonlabor conditions and also the presence of two novel TIMP family members (TIMP-3 and TIMP-4). METHOD OF STUDY: Amniochorionic membranes collected from women undergoing Cesarean section and were cultured in an organ explant system. Membranes were also collected from laboring women after vaginal delivery. Samples were subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) using primers specific for TIMP-1, TIMP-2, TIMP-3, and TIMP-4. Localization of TIMP mRNAs was accomplished by in situ hybridization, and peptides were localized by immunocytochemistry. RESULTS: RT-PCR data demonstrated the expression of all the TIMPs in tissues from laboring and nonlaboring women as well as in cultured membranes. TIMP-4 expression was seen in RT-PCR, however, only a faint band was visible in all the tissues tested. In situ hybridization localized the TIMP mRNAs to the amnion, chorion, and to scattered cells in the connective tissue. CONCLUSION: Human fetal membrane cells (amniochorion and decidua) express mRNA for all the TIMPs studied so far.

Extraembryonic Membranes↗
PubMed1998

Expression of a progelatinase activator (MT1-MMP) in human fetal membranes.

PROBLEM: The finding of MMP-2 (which degrades type IV collagen) and TIMP-2 (the tissue inhibitor of MMP) in fetal membranes suggests the possibility of membrane self-destruction as an etiology of premature rupture of fetal membranes. MMP-2 is activated by a membrane-bound MMP (MT1-MMP). This study was undertaken to detect the presence of MT1-MMP in human fetal membranes. METHOD OF STUDY: Fetal membranes were placed in an organ explant system and stimulated with lipopolysaccaride (LPS). MT1-MMP expression was studied in frozen tissues by reverse transcriptase (RT)-polymerase chain reaction (PCR) using primers designed in our laboratory. DNA sequence analysis was performed to verify the specificity of PCR products. In situ hybridization and immunocytochemistry were used to localize MT1-MMP mRNA and peptide, respectively. RESULTS: RT-PCR data indicated the presence of mRNA for MT1-MMP in fetal membranes. Although PCR is not quantitative, no differences in mRNA band intensities were noticed after LPS stimulation. MT1-MMP expression was constitutive throughout the culture period. In situ hybridization demonstrated amnion, chorionic laeve, cytotrophoblast cells, and the cells in the reticular and spongy layer of the extracellular matrix as the origin of MT1-MMP mRNA and peptide. CONCLUSIONS: This is the first study documenting the amniochorionic membrane as a source of MT1-MMP mRNA and peptide. Activation of progelatinase A requires the presence of this membrane-associated MMP. The finding of MT1-MMP in a tissue already known to produce MMP-2 and TIMP-2 documents the full system for activation and inhibition of this gelatinase. During infection, an imbalance in the expression of MT1-MMP, MMP-2 and TIMP-2 may constitute an endogenous pathway of membrane degradation.

Enzyme Precursors↗
PubMed1998

Immunoreactivity of human fetal membranes to peptidoglycan polysaccharide (PGPS): cytokine response.

OBJECTIVE: Group-B Streptococcus has been associated with preterm labor and other pregnancy related complications. This study was performed to evaluate the effect of peptidoglycan polysaccharide (PGPS) derived from a beta hemolytic Streptococcal cell wall on amniochorion cytokine production and to compare PGPS effects with lipopolysaccharide (LPS), which is the Gram negative counterpart of PGPS. STUDY DESIGN: Amniochorionic membranes collected from women not in labor, and undergoing elective repeat C-section were placed in an organ explant system. Membranes were stimulated separately with 50 ng/ml of small (100p), large (10s) fractions of PGPS or LPS respectively immediately after collection and after a stabilization period of 48 hrs. Media samples were collected at 3, 6, 9, 12 and 24 hrs for protein analysis after each stimulation. Media samples were analyzed by ELISA for IL-6 and IL-8. RESULTS: Both forms of PGPS and LPS stimulated IL-6 and IL-8 production by human fetal membranes. Of note is that LPS stimulated IL-6 to a greater degree than IL-8, while PGPS stimulated IL-8 to a greater degree than IL-6. No statistical difference was seen in the levels of either one of these cytokines for the larger or smaller fragments of PGPS. Time course studies documented a 3-hour lag phase when tissues are stimulated directly after collection which was absent when tissues are stimulated after a 48-hour stabilization period. CONCLUSION: Both PGPS and LPS stimulate cytokine production differently from fetal membranes. This supports the theory that different bacteria may affect the host in contrasting ways which may lead to a distinct host response, i.e. PROM vs. PTL.

Amnion↗
PubMed1997

Collagenolytic enzymes (gelatinases) and their inhibitors in human amniochorionic membrane.

OBJECTIVE: This study was designed to investigate the presence of matrix metalloproteinase-2 (gelatinase A), matrix metalloproteinase-9 (gelatinase B), and their natural inhibitors in both cultured amniochorionic membrane and membrane obtained from women with infection-associated preterm labor. STUDY DESIGN: Amniochorionic membranes were collected from women with documented intraamniotic infection and from women not in labor undergoing elective repeat cesarean section with no signs of infection or other complications of pregnancy. Normal membranes were cultured and exposed to endotoxin and peptidoglycan polysaccharide. Messenger ribonucleic acid expression for gelatinase A, gelatinase B, and tissue inhibitors of matrix metalloproteinase types 1 and 2 was studied with use of reverse transcriptase-polymerase chain reaction and localization of messenger ribonucleic acid was accomplished with use of in situ hybridization. Release of gelatinases from the membranes was studied with gelatin zymography. Tissue inhibitors of matrix metalloproteinase peptides were localized with use of immunocytochemistry. RESULTS: The expression of matrix metalloproteinase types 2 and 9 was seen in amniochorionic membranes in culture. Matrix metalloproteinase-2 was seen in membranes from nonlaboring women and in women with intraamniotic infection, whereas matrix metalloproteinase-9 was seen only in membranes from women with intraamniotic infection. The matrix metalloproteinase-9 expression could also be induced by lipopolysaccharide or peptidoglycan polysaccharide stimulation in culture. In situ hybridization localized messenger ribonucleic acid for these matrix metalloproteinases to both amnion and chorion. Zymogram studies showed the activity of matrix metalloproteinase-2 in normal resting membrane and cultured membrane. Matrix metalloproteinase-9 was induced by culture conditions. Tissue inhibitor of matrix metalloproteinase-1 and tissue inhibitor of matrix metalloproteinase-2 messenger ribonucleic acid was seen in normal, infected, and cultured membranes. In situ hybridization data indicated that these messages were mainly produced by chorion, but they were also seen in amnion. Immunohistochemistry demonstrated the presence of tissue inhibitor of matrix metalloproteinase-1 and tissue inhibitor of matrix metalloproteinase-2 peptides in both amnion and chorion and in cells of the reticular layer of the matrix. CONCLUSION: Normal amniochorionic membrane is a source of matrix metalloproteinase-2 and tissue inhibitors of matrix metalloproteinases. Culture conditions and infection induce matrix metalloproteinase-9 expression and release from amniochorion. These findings suggest that these collagenolytic enzymes may play a role in premature rupture of the membranes in infection, which can lead to preterm labor.

Amnion↗
PubMed1997

Interleukin-10 and transforming growth factor-beta inhibit amniochorion tumor necrosis factor-alpha production by contrasting mechanisms of action: therapeutic implications in prematurity.

OBJECTIVE: This study was designed to detect the regulatory effect of the immunoinhibitory cytokines interleukin-10 and transforming growth factor-beta on the amniochorion production of tumor necrosis factor-alpha. STUDY DESIGN: Amniochorionic membranes were collected from women undergoing elective repeat cesarean section with no history of infection. Membranes were placed in organ explant culture for 48 hours and then stimulated with lipopolysaccharide (50 ng/ml), lipopolysaccharide plus interleukin-10 (50/ 50, 50/100 ng/ml), interleukin-10 (50 and 100 ng/ml), lipopolysaccharide plus transforming growth factor-beta (50/50 and 50/100 ng/ml), and transforming growth factor-beta (50 and 100 ng/ml). At the end of a 24-hour stimulation tissue samples were frozen for ribonucleic acid analysis and media samples were frozen for enzyme-linked immunosorbent assay. Quantitation of the messenger ribonucleic acid was accomplished by quantitative competitive polymerase chain reaction, and tumor necrosis factor-alpha protein was assayed by use of enzyme-linked immunosorbent assay. RESULTS: Lipopolysaccharide stimulation of fetal membranes produced approximately 60,000 molecules of tumor necrosis factor-alpha messenger ribonucleic acid, whereas control tissue produced none. Lipopolysaccharide plus interleukin-10 stimulation resulted in a dose-dependent decrease in tumor necrosis factor-alpha messenger ribonucleic production (transcriptional regulation) to 6000 (50/50) and 600 (50/100) molecules. Enzyme-linked immunosorbent assay performed on media samples from these experiments demonstrated a dose-dependent reduction in tumor necrosis factor-alpha peptide release. Stimulation of membranes with lipopolysaccharide plus transforming growth factor-beta had minimal effects on tumor necrosis factor-alpha messenger ribonucleic acid and protein production compared with lipopolysaccharide-treated samples. Membranes stimulated with interleukin-10 alone showed no effect on messenger ribonucleic acid or protein levels and remained similar to the levels seen in control tissues. In the absence of lipopolysaccharide, transforming growth factor-beta treatment produced a dramatic decrease in tumor necrosis factor-alpha peptide levels without affecting messenger ribonucleic acid levels. CONCLUSION: In the presence of a stimulatory agent, interleukin-10 down-regulates tumor necrosis factor-alpha release from cultured human amniochorionic membranes. Transforming growth factor-beta seems to have some stimulatory effect on transcription, and no effect on translation was seen with concurrent lipopolysaccharide stimulation. However, down-regulation of tumor necrosis factor-alpha peptide by transforming growth factor was seen in fetal membranes when not overridden by an inflammatory stimulant. This study suggests that interleukin-10 and transforming growth factor-beta can regulate tumor necrosis factor-alpha release from amniochorion under different conditions and by a different mechanism.

Amnion↗
PubMed1997

Sequence-specific Priming and Exonuclease-released Fluorescence Assay for Rapid and Reliable HLA-A Molecular Typing.

Background: The rapid and sensitive method described for low-resolution DNA typing of alleles at the HLA-A locus is based on a sequence-specific primer polymerase chain reaction approach that eliminates post-polymerase chain reaction gel electrophoresis to analyze the results. Methods and Results: This method takes advantage of the 5' --> 3' exonuclease activity of the Taq polymerase normally present during polymerase chain reaction. This sequence-specific priming and exonuclease-released fluorescence assay was conducted on 40 DNA samples derived from homozygous cell lines and peripheral blood lymphocytes that represented the majority of the HLA-A alleles. There was 100% condordance with the results obtained by serologic typing of these same samples. Conclusions: The advantages of this system are that assays can be performed in 2 hours, the results are easy to interpret, and no gel electrophoresis is required. The same strategy can be used for high-resolution DNA typing once appropriate primer pairs are added to the panel used for first-level discrimination.

Journal Article↗
PubMed1997

Double-labeled fluorescent probes for 5' nuclease assays: purification and performance evaluation.

An inexpensive method for the purification and evaluation of user-synthesized or crude commercially prepared double-labeled fluorescent probes is presented. These probes exhibit the characteristics required for use in 5'-nuclease assays, including efficient reporter dye quenching, target specificity and susceptibility to cleavage by Taq DNA polymerase during PCR amplification. The method is suitable for research laboratories that wish to develop 5' nuclease assays for the detection of PCR-amplified target sequences to eliminate the requirement for agarose gels and to advance throughput.

5'-Nucleotidase↗
PubMed1996

Exploratory factor analysis of MRI brain structure measures in schizophrenia.

Much of the literature shows various regional structural brain abnormalities in schizophrenia, but the complexity and variability of brain makes it difficult to determine how these regions are related. Statistical methods which estimate factors underlying patterns of covariance have not been widely used, but could be useful for analyzing such complex data. We applied exploratory and confirmatory factor analysis procedures to specific cortical and subcortical regional brain volume measures from MRI data in 60 normal and 44 schizophrenic subjects. Basal ganglia, heteromodal cortical gray, and medial temporal lobe factors were present in both the normal and the schizophrenia groups. The factor structure observed in the normal group showed a high degree of bilateral symmetry which is present but disrupted in the schizophrenia group. In the bilateral data, the disruption is most pronounced with medial and lateral temporal lobe structures including entorhinal cortex and anterior and posterior superior temporal gyri. There was a significant correlation between the basal ganglia factor and the heteromodal cortical gray factor in the normal group that was not present in the schizophrenia group. In the unilateral data, left posterior superior temporal gyrus did not load onto any factor in the schizophrenia group. Confirmatory factor analyses showed significant differences between the two groups in factor structure. A number of specific brain regions are affected in schizophrenia, and structural relationships between groups of regions also are abnormal. The results suggest that heteromodal dorsolateral prefrontal and superior temporal cortical gray regions are structurally related, whereas inferior parietal cortical gray is less so. These results should be viewed as preliminary as the ratio of parameters to subjects was relatively low, and replication is needed. However, the results demonstrate the potential utility of latent structure methods such as factor analysis in study of complex relationships in neuropsychiatric data.

Adult↗
PubMed1996

Inflammatory cytokine (interleukins 1, 6 and 8 and tumor necrosis factor-alpha) release from cultured human fetal membranes in response to endotoxic lipopolysaccharide mirrors amniotic fluid concentrations.

OBJECTIVE: This study was conducted to quantitate and compare the amount of cytokines released from human fetal membranes in response to treatment with bacterial lipopolysaccharide and to compare this with amniotic fluid levels. STUDY DESIGN: Amniochorionic membranes were collected from women undergoing elective repeat cesarean section and showing no signs of infection- or pregnancy-related complications. Membranes were maintained in an organ explant system and stimulated with bacterial lipopolysaccharide for 24 hours. Media samples were collected and stored at -20 degrees C until cytokine levels were assayed by enzyme-linked immunosorbent assay. RESULTS: Enzyme-linked immunosorbent assay results demonstrated that lipopolysaccharide stimulated production of interleukins 1, 6 and 8 and tumor necrosis factor-alpha by the fetal membranes in comparison with the control cultures. A greater release of interleukin-6 and interleukin-8 compared with interleukin-1 and tumor necrosis factor-alpha was noticed. The relationships between cytokine concentrations observed in culture mirror those seen in amniotic fluid. CONCLUSION: Amniochorionic membranes can respond to an infectious process with increased secretion of interleukins 1, 6 and 8 and tumor necrosis factor-alpha. Cytokines produced from both amnion and chorion (interleukin-6 and interleukin-8) are released in greater quantities than those cytokines produced from chorion or amnion alone (interleukin-1 and tumor necrosis factor-alpha). These studies support a major role for amnion in infection-induced preterm labor.

Amniotic Fluid↗