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Biomedical subjects

X Qin

Publications and source records attributed to X Qin.

At least 19 recordsLinked to original sources

S-RNase uptake by compatible pollen tubes in gametophytic self-incompatibility.

Many flowering plants avoid inbreeding through a genetic mechanism termed self-incompatibility. An extremely polymorphic S-locus controls the gametophytic self-incompatibility system that causes pollen rejection (that is, active arrest of pollen tube growth inside the style) when an S-allele carried by haploid pollen matches one of the S-alleles present in the diploid style. The only known product of the S-locus is an S-RNase expressed in the mature style. The pollen component to this cell-cell recognition system is unknown and current models propose that it either acts as a gatekeeper allowing only its cognate S-RNase to enter the pollen tube, or as an inhibitor of non-cognate S-RNases. In the latter case, all S-RNases are presumed to enter pollen tubes; thus, the two models make diametrically opposed predictions concerning the entry of S-RNases into compatible pollen. Here we use immunocytochemical labelling of pollen tubes growing in styles to show accumulation of an S-RNase in the cytoplasm of all pollen-tube haplotypes, thus providing experimental support for the inhibitor model.

Pollen↗

Identification and functional characterization of a new gene encoding the mouse terminal complement inhibitor CD59.

CD59 is a 18- to 20-kDa, GPI-anchored membrane protein that functions as a key regulator of the terminal step of the complement activation cascade. It restricts binding of C9 to the C5b-8 complex, thereby preventing the formation of the membrane attack complex (C5b-9 of complement). A single human CD59 gene has been identified, and corresponding genetic homologues from rat, mouse, and pig have been characterized in previous studies. In this study, we report the discovery and functional characterization of a separate cd59 gene in the mouse (referred to as cd59b, the previously characterized mouse cd59 gene as cd59a). Mouse cd59b is 85% and 63% identical to cd59a at the nucleotide and amino acid level, respectively. In cDNA transfection experiments with Chinese hamster ovary cells, peptide-tagged cd59b was detected on the cell surface by flow cytometry and was shown to be susceptible to phosphatidylinositol-specific phospholipase C cleavage. Chinese hamster ovary cells expressing cd59b were significantly more resistant than control cells to human and mouse complement-mediated lysis. These results suggest that cd59b encodes a GPI-anchored protein that is functionally active as a membrane attack complex inhibitor. Northern blot analysis revealed that cd59b is expressed selectively in the mouse testis. In contrast, the major transcript of cd59a was shown to be expressed at high levels in the heart, kidney, liver, and lung, but only minimally in the testis. These results revealed the existence of two distinct cd59 genes in the mouse that are differentially regulated and that may have nonoverlapping physiological functions in vivo.

Amino Acid Sequence↗

Evidence that the interaction between insulin-like growth factor (IGF)-II and IGF binding protein (IGFBP)-4 is essential for the action of the IGF-II-dependent IGFBP-4 protease.

A variety of human cell types, including human osteoblasts (hOBs), produce an IGFBP-4 protease, which cleaves IGFBP-4 in the presence of IGF-II. Recently, the pregnancy-associated plasma protein (PAPP)-A has been determined to be the IGF-II-dependent IGFBP-4 protease produced by human fibroblasts. This study sought to define the mechanism by which IGF-II enhances IGFBP-4 proteolysis. Addition of PAPP-A antibody blocked the IGFBP-4 proteolytic activity in hOB conditioned medium (CM), suggesting that PAPP-A is the major IGFBP-4 protease in hOB CM. Pre-incubation of IGFBP-4 with IGF-II, followed by removal of unbound IGF-II, led to IGFBP-4 proteolysis without further requirement of the presence of IGF-II in the reaction. In contrast, prior incubation of the partially purified IGFBP-4 protease from either hOB CM or human pregnancy serum with IGF-II did not lead to IGFBP-4 proteolysis unless IGF-II was re-added to the assays. To further confirm that the interaction between IGF-II and IGFBP-4 is required for IGFBP-4 protease activity, we prepared IGFBP-4 mutants, which contained the intact cleavage site (Met135-Lys136) but lacked the IGF binding activity, by deleting the residues Leu72-His74 in the IGF binding domain or Cys183-Glu237 that contained an IGF binding enhancing motif. The IGFBP-4 protease was unable to cleave these IGFBP-4 mutants, regardless of whether or not IGF-II was present in the assay. Conversely, an IGFBP-4 mutant with His74 replaced by an Ala, which exhibited normal IGF binding activity, was effectively cleaved in the presence of IGF-II. Taken together, these findings provided strong evidence that the interaction between IGF-II and IGFBP-4, rather than the direct interaction between IGF-II and IGFBP-4 protease, is required for optimal IGFBP-4 proteolysis.

Antibodies↗

Determination of neurotoxin 3-N-oxalyl-2,3-diaminopropionic acid and non-protein amino acids in Lathyrus sativus by precolumn derivatization with 1-fluoro-2,4-dinitrobenzene.

A rapid and simple method is presented for determining neuro-excitatory nonprotein amino acid 3-N-oxalyl-2,3-diaminopropionic acid (beta-ODAP) and non-protein amino acids in Lathyrus sativus. Seed and foliage extracts of Lathyrus sativus were treated with 1-fluoro-2,4-dinitrobenzene (FDNB) and a reversed-phase high-performance liquid chromatography method (RP HPLC) for the separation of the derivatives in the pmol range is reported. The RP HPLC method and a colorimetric method were compared for measuring ODAP.

Amino Acids↗

16S rDNA sequence analysis of Xylella fastidiosa strains.

The 16S rDNA encoding the small subunit ribosomal RNA were amplified by PCR, cloned, and sequenced from 16 strains of Xylella fastidiosa originating from nine different hosts. In pair-wise comparisons, X. fastidiosa strains showed a maximum variation of 1.0% or 14 nucleotide positions. When all 16 sequences were considered as a set, 54 variable positions were found. Analysis of the sequence data indicated that the X. fastdiosa strains formed three rDNA groups. Group one includes Pierce's disease and mulberry leaf scorch strains; Group two, periwinkle wilt, plum leaf scald, phony peach, oak leaf scorch, and elm leaf scorch strains; and Group three, citrus variegated chlorosis and coffee leaf scorch strains. All X. fastidiosa strains exhibited significantly higher levels of sequence heterogeneity (63 to 83 nucleotide positions) when compared to species from Xanthomonas and Stenotrophomonas. Our data demonstrate that 16S rDNA sequence data could provide valuable information for future classification of X. fastidiosa at the sub-species level.

DNA, Ribosomal↗

Heterozygous DNA mismatch repair gene PMS2-knockout mice are susceptible to intestinal tumor induction with N-methyl-N-nitrosourea.

PMS2-deficient (PMS2(-/-)) mice are hypersensitive to N-methyl-N-nitrosourea (MNU)-induced thymic lymphomas based on the failure to initiate mismatch repair (MMR) at O(6)-methylguanine:T mismatches formed after MNU exposure. However, heterozygous PMS2 knockout (PMS2(+/-)) mice do not develop spontaneous tumors, suggesting that they have sufficient MMR function to prevent genomic instability. We hypothesized that in PMS2(+/-) mice, exogenous carcinogens may either mutationally knockout the remaining normal allele leading cells to develop tumors or introduce sufficient DNA adducts and mismatches to overload the lower capacity for MMR, leading in either case to an increased rate of tumor production. In the present study, PMS2(+/-) mice and their littermate PMS2(+/+) mice were monitored for tumor incidence following MNU treatment. Mice were given 50 mg MNU/kg i.p. when 5 weeks old. They demonstrated a similar incidence of thymic lymphomas, suggesting that expression of the single normal PMS2 allele is sufficient to protect the thymus and implying that a single dose of MNU may not efficiently knock out the remaining PMS2 allele in the thymus. Surprisingly, PMS2(+/-) mice were significantly more likely to develop intestinal tumors-both adenomas and adenocarcinomas-after MNU than were PMS2(+/+) mice (2.34 +/- 0.34 tumors per mouse versus 1.34 +/- 0.25 tumors per mouse; P < 0.05). The intestinal tumors were located mainly in the small intestine. However, these tumors in both the PMS2(+/-) mice and PMS2(+/+) mice did not show microsatellite instability characteristic of loss of MMR. These results suggest that a single normal PMS2 allele can protect thymus but not intestine from MNU carcinogenesis. Organ-specific factors might influence MMR- mediated resistance to methylating agents. Heterozygous PMS2 knockout mice may be used as a promising animal model for intestinal tumorigenesis studies involving environmental carcinogens.

Adenosine Triphosphatases↗

Protection against malignant progression of spontaneously developing liver tumors in transgenic mice expressing O(6)-methylguanine-DNA methyltransferase.

To study the effect of O(6)-methylguanine-DNA methyltransferase (MGMT) on carcinogenesis, we have previously generated MGMT transgenic mice overexpressing the bacterial MGMT gene, ada, and demonstrated that high MGMT levels in the liver suppress induction of liver tumors after treatment with an alkylating hepatocarcinogen. To examine the effects of life-long elevation of MGMT activity on mouse spontaneous liver tumor development, ada-transgenic and control non-transgenic mice were compared. We also examined mutations at codon 61 of the H-ras oncogene, reported as a hot spot in mouse liver tumors, using a direct DNA sequencing method. The results revealed no significant difference in tumor incidence or mutation spectrum, but interestingly, ada-transgenic mice were found to have fewer malignant tumors and survived longer, indicating a possible protective role of MGMT against malignant conversion.

Animals↗

Analysis of a gene cluster of Enterococcus faecalis involved in polysaccharide biosynthesis.

Previously, we described a gene cluster of Enterococcus faecalis OG1RF that produced an antigenic polysaccharide when cloned in Escherichia coli. The polysaccharide antigen was not detectable in E. faecalis strains, however. Here, we show by reverse transcriptase-PCR that the 16 genes in this region are transcribed in OG1RF. Gene disruption of orfde4, encoding a putative glycosyl transferase, and orfde6, a putative dTDP-rhamnose biosynthesis gene, generated two OG1RF mutants. The mutants showed delayed killing and a higher 50% lethal dose in a mouse peritonitis model. In addition, two mucoid E. faecalis isolates from patients with chronic urinary tract infections were found to produce the polysaccharide antigen.

Animals↗

Effects of Enterococcus faecalis fsr genes on production of gelatinase and a serine protease and virulence.

Three agr-like genes (fsrA, fsrB, and fsrC, for Enterococcus faecalis regulator) were found upstream of the previously reported gelatinase gene (gelE) and a downstream putative serine protease gene (sprE; accession number Z12296) of Enterococcus faecalis OG1RF. The deduced amino acid sequence of fsrA shows 26% identity and 38% similarity to Staphylococcus aureus AgrA (the response regulator of the accessory gene regulator system in the agr locus), FsrB shows 23% identity and 41% similarity to S. aureus AgrB, and FsrC shows 23% identity and 36% similarity to S. aureus AgrC (the sensor transducer of Agr system). Northern blot analysis suggested that gelE and sprE are cotranscribed and that fsrB and fsrC are also cotranscribed in OG1RF. Northern blot analysis of fsrA, fsrB, fsrC, gelE, and sprE insertion mutants showed that fsrB, fsrC, gelE, and sprE are not expressed in fsrA, fsrB, and fsrC mutants, while insertion in an open reading frame further upstream of fsrA did not effect the expression of these genes, suggesting that agr-like genes may be autoregulated and that they regulate gelE and sprE expression, as further confirmed by complementation of fsr gene mutations with a 6-kb fragment which contains all three fsr genes in the shuttle vector, pAT18. Testing of 95 other isolates of E. faecalis showed that 62% produced gelatinase (Gel(+)), while 91% (including all Gel(+) strains) hybridized to a gelE probe; 71% (including all Gel(+) strains) hybridized to an fsr probe, corroborating the conclusion that both gelE and fsr are necessary for gelatinase production. Testing of fsrA, fsrB, and sprE mutants in a mouse peritonitis model showed that sprE and agr-like gene mutants resulted in highly significantly prolonged survival compared to the parent strain OG1RF, a finding similar to what we had previously shown for a gelE mutant. These results suggest that sprE and agr-like genes contribute to the virulence of E. faecalis OG1RF in this model.

Amino Acid Sequence↗

Enterococcus faecalis adhesin, ace, mediates attachment to extracellular matrix proteins collagen type IV and laminin as well as collagen type I.

Adhesin-mediated binding to extracellular matrix (ECM) proteins is thought to be a crucial step in the pathogenic process of many bacterial infections. We have previously reported conditional adherence of most Enterococcus faecalis isolates, after growth at 46 degrees C, to ECM proteins collagen types I and IV and laminin; identified an E. faecalis-specific gene, ace, whose encoded protein has characteristics of a bacterial adhesin; and implicated Ace in binding to collagen type I. In this study, we constructed an ace disruption mutant from E. faecalis strain OG1RF that showed marked reduction in adherence to collagen types I and IV and laminin when compared to the parental OG1RF strain after growth at 46 degrees C. Polyclonal immune serum raised against the OG1RF-derived recombinant Ace A domain reacted with a single approximately 105-kDa band of mutanolysin extracts from OG1RF grown at 46 degrees C, while no band was detected in extracts from OG1RF grown at 37 degrees C, nor from the OG1RF ace mutant grown at 37 or 46 degrees C. IgGs purified from the anti-Ace A immune serum inhibited adherence of 46 degrees C-grown E. faecalis OG1RF to immobilized collagen type IV and laminin as well as collagen type I, at a concentration as low as 1 microg/ml, and also inhibited the 46 degrees C-evoked adherence of two clinical isolates tested. We also showed in vitro interaction of collagen type IV with Ace from OG1RF mutanolysin extracts on a far-Western blot. Binding of recombinant Ace A to immobilized collagen types I and IV and laminin was demonstrated in an enzyme-linked immunosorbent assay and was shown to be concentration dependent. These results indicate that Ace A mediates the conditional binding of E. faecalis OG1RF to collagen type IV and laminin in addition to collagen type I.

Animals↗

Comparison of Escherichia coli O157:H7 antigen detection in stool and broth cultures to that in sorbitol-MacConkey agar stool cultures.

We evaluated the Meridian IC-STAT direct fecal and broth culture antigen detection methods with samples from children infected with Escherichia coli O157:H7 and correlated the antigen detection results with the culture results. Stools of 16 children who had recently had stool cultures positive for this pathogen (population A) and 102 children with diarrhea of unknown cause (population B) were tested with the IC-STAT device (direct testing). Fecal broth cultures were also tested with this device (broth testing). The results were correlated to a standard of the combined yield from direct culture of stools on sorbitol-MacConkey (SMAC) agar and culture of broth on SMAC agar. Eleven (69%) of the population A stool specimens yielded E. coli O157:H7 when plated directly on SMAC agar. Two more specimens yielded this pathogen when the broth culture was similarly plated. Of these 13 stool specimens, 8 and 13 were positive by direct and broth testing (respective sensitivities, 62 and 100%). Compared to the sensitivity of a simultaneously performed SMAC agar culture, the sensitivity of direct testing was 73%. Three (3%) of the population B stool specimens contained E. coli O157:H7 on SMAC agar culture; one and three of these stool specimens were positive by direct and broth testing, respectively. The direct and broth IC-STAT tests were 100% specific with samples from children from population B. Direct IC-STAT testing of stools is rapid, easily performed, and specific but is insufficiently sensitive to exclude the possibility of infection with E. coli O157:H7. Performing the IC-STAT test with a broth culture increases its sensitivity. However, attempts to recover E. coli O157:H7 by culture should not be abandoned but, rather, should be increased when the IC-STAT test result is positive.

Antigens, Bacterial↗

Studies on human pregnancy-induced insulin-like growth factor (IGF)-binding protein-4 proteases in serum: determination of IGF-II dependency and localization of cleavage site.

Insulin-like growth factor (IGF)-binding protein-4 (IGFBP-4), a consistent inhibitor of IGF action, is subject to proteolytic cleavage by the IGF-II-dependent IGFBP-4 protease. However, regulation of the IGF-II-dependent IGFBP-4 protease in vivo is not known. As IGFBP proteases are known to be triggered during pregnancy, we systematically evaluated the changes in IGFBP-4 proteolysis by serum collected throughout human pregnancy. Results from in vitro protease assays using recombinant IGFBP-4 revealed that IGFBP-4 proteolysis determined in both the presence and absence of exogenous IGF-II significantly increased during the first and second trimesters and reached a plateau by the third trimester. However, in the absence of IGF-II, IGFBP-4 proteolysis by pregnancy serum was only observed after prolonged incubation. IGF-II dose dependently increased IGFBP-4 proteolysis by pregnancy serum, with maximal stimulation observed at a concentration of 0.7 mol/L relative to IGFBP-4. In contrast, IGF-II at an equimolar dose had little effect on proteolysis of recombinant human IGFBP-3, whereas excess IGF-II reproducibly inhibited recombinant human IGFBP-3 proteolysis by pregnancy serum. Although IGF-II enhanced IGFBP-4 proteolysis, results from N-terminal sequence and mass spectrometric analyses of IGFBP-4 proteolytic fragments demonstrate that the cleavage site (Met135-Lys136) in human IGFBP-4 was not altered by IGF-II. Deletion of the residues 121-141 containing this cleavage site blocked IGFBP-4 proteolysis. These findings demonstrate that the increase in IGFBP-4 proteolysis during pregnancy was accounted for mainly by the IGF-II-dependent IGFBP-4 proteolysis. Because IGFBP-4 is a potent inhibitor of IGF actions, it can be speculated that the pregnancy-induced IGFBP-4 proteases may play an important role in regulating fetal growth.

Adult↗

Brain expression of presenilins in sporadic and early-onset, familial Alzheimer's disease.

BACKGROUND: Mutations in the presenilin proteins cause early-onset, familial Alzheimer's disease (FAD). MATERIALS AND METHODS: We characterized the cellular localization and endoproteolysis of presenilin 2 (PS2) and presenilin 1 (PS1) in brains from 25 individuals with presenilin-mutations causing FAD, as well as neurologically normal individuals and individuals with sporadic Alzheimer's disease (AD). RESULTS: Amino-terminal antibodies to both presenilins predominantly decorated large neurons. Regional differences between the broad distributions of the two presenilins were greatest in the cerebellum, where most Purkinje cells showed high levels of only PS2 immunoreactivity. PS2 endoproteolysis in brain yielded multiple amino-terminal fragments similar in size to the PS1 amino-terminal fragments detected in brain. In addition, two different PS2 amino-terminal antibodies also detected a prominent 42 kDa band that may represent a novel PS2 form in human brain. Similar to PS1 findings, neither amino-terminal nor antiloop PS2 antibodies revealed substantial full-length PS2 in brain. Immunocytochemical examination of brains from individuals with the N141I PS2 mutation or eight different PS1 mutations, spanning the molecule from the second transmembrane domain to the large cytoplasmic loop domain, revealed immunodecoration of no senile plaques and only neurofibrillary tangles in the M139I PS1 mutation stained with PS1 antibodies. CONCLUSIONS: Overall presenilin expression and the relative abundance of full-length and amino-terminal fragments in presenilin FAD cases were similar to control cases and sporadic AD cases. Thus, accumulation of full-length protein or other gross mismetabolism of neither PS2 nor PS1 is a consequence of the FAD mutations examined.

Age of Onset↗

The 9-cis-epoxycarotenoid cleavage reaction is the key regulatory step of abscisic acid biosynthesis in water-stressed bean.

Abscisic acid (ABA), a cleavage product of carotenoids, is involved in stress responses in plants. A well known response of plants to water stress is accumulation of ABA, which is caused by de novo synthesis. The limiting step of ABA biosynthesis in plants is presumably the cleavage of 9-cis-epoxycarotenoids, the first committed step of ABA biosynthesis. This step generates the C(15) intermediate xanthoxin and C(25)-apocarotenoids. A cDNA, PvNCED1, was cloned from wilted bean (Phaseolus vulgaris L.) leaves. The 2, 398-bp full-length PvNCED1 has an ORF of 615 aa and encodes a 68-kDa protein. The PvNCED1 protein is imported into chloroplasts, where it is associated with the thylakoids. The recombinant protein PvNCED1 catalyzes the cleavage of 9-cis-violaxanthin and 9'-cis-neoxanthin, so that the enzyme is referred to as 9-cis-epoxycarotenoid dioxygenase. When detached bean leaves were water stressed, ABA accumulation was preceded by large increases in PvNCED1 mRNA and protein levels. Conversely, rehydration of stressed leaves caused a rapid decrease in PvNCED1 mRNA, protein, and ABA levels. In bean roots, a similar correlation among PvNCED1 mRNA, protein, and ABA levels was observed. However, the ABA content was much less than in leaves, presumably because of the much smaller carotenoid precursor pool in roots than in leaves. At 7 degrees C, PvNCED1 mRNA and ABA were slowly induced by water stress, but, at 2 degrees C, neither accumulated. The results provide evidence that drought-induced ABA biosynthesis is regulated by the 9-cis-epoxycarotenoid cleavage reaction and that this reaction takes place in the thylakoids, where the carotenoid substrate is located.

Abscisic Acid↗

Mice defective in the DNA mismatch gene PMS2 are hypersensitive to MNU induced thymic lymphoma and are partially protected by transgenic expression of human MGMT.

DNA mismatch repair (MMR) stabilizes the cellular genome. Mice defective in the MMR gene PMS2 are susceptible to spontaneous thymic lymphoma and sarcomas. To determine the sensitivity of PMS2 knockout mice to environmental carcinogens and the protective effect of O6-methylguanine DNA methyltransferase (MGMT), heterozygous PMS2 knockout mice and human MGMT (hMGMT) transgenic mice were mated and the PMS2-/- and PMS2+/+ with or without hMGMT offspring were treated at 5 weeks of age with 50 mg/kg N-methyl-N-nitrosourea (MNU). MNU produces carcinogenic O6-methylguanine (O6-meG) adducts, resulting in thymic lymphoma in mice, which can be prevented in normal mice by overexpression of hMGMT. A significantly higher incidence of thymic lymphomas was observed in MNU-treated PMS2-/- mice, compared to wildtype PMS2+/+ mice (100 vs 52%; P < 0.001). The mean latency of lymphomas was also significantly shortened in PMS2-/- mice (81 vs 102 days, P < 0.01). Transgenic expression of hMGMT significantly but incompletely blocked MNU lymphomagenesis in PMS2-/- mice. The incidence of lymphomas in PMS2-/-/hMGMT+ mice was reduced to 80% (P < 0.01) and mean latency increased to 91 days (P < 0.05). Thymic lymphomagenesis was efficiently blocked in PMS2+/+/hMGMT+ mice with rapid repair of O6-meG. Since O6-meG:T mismatches in MMR+ cells may trigger mismatch repair resulting in abortive repair and cell death whereas in the absence of MMR, these mismatches are converted to A:T, we predicted that G to A point mutations in codon 12 of the K-ras gene would occur. In this study, we found G to A point mutations in codon 12 of the K-ras gene in many tumors. Thus, in MMR deficient tissues, methylating agents induce point mutations in cells with a higher rate of cell survival which together are potently carcinogenic in the thymus. These data suggest that PMS2 defective lymphomas may arise by the concerted action of environmental and perhaps endogenous methylation of DNA coupled to genomic instability.

Adenosine Triphosphatases↗

Matrix metalloproteinase deficiencies affect contact hypersensitivity: stromelysin-1 deficiency prevents the response and gelatinase B deficiency prolongs the response.

Matrix metalloproteinases (MMPs) are expressed by T cells and macrophages, but there is a paucity of evidence for their role in immune responses. We have studied mice with deficiencies of stromelysin-1 (MMP-3) or gelatinase B (MMP-9) in a dinitrofluorobenzene (DNFB)-induced model of contact hypersensitivity (CHS). Stromelysin-1-deficient mice showed a markedly impaired CHS response to topical DNFB, although they responded normally to cutaneously applied phenol, an acute irritant. Lymphocytes from lymph nodes of DNFB-sensitized stromelysin-1-deficient mice did not proliferate in response to specific soluble antigen dinitrobenzenesulfonic acid, but did proliferate identically to lymph node lymphocytes from wild-type mice when presented with the mitogen Con A. An intradermal injection of stromelysin-1 immediately before DNFB sensitization rescued the impaired CHS response to DNFB in stromelysin-1-deficient mice. Unlike stromelysin-1-deficient mice, gelatinase B-deficient mice exhibited a CHS response comparable to wild-type controls at 1 day postchallenge, but the response persisted beyond 7 days in contrast to the complete resolution observed in wild-type mice by 7 days. However, gelatinase B-deficient mice had a normal rate of resolution of acute inflammation elicited by cutaneous phenol. Gelatinase B-deficient mice failed to show IL-10 production at the site of CHS, an essential feature of resolution in control mice. These results indicate that stromelysin-1 and gelatinase B serve important functions in CHS. Stromelysin-1 is required for initiation of the response, whereas gelatinase B plays a critical role in its resolution.

Animals↗

Intestinal synthesis and lymphatic secretion of apolipoprotein A-IV after cessation of duodenal fat infusion: mediation by bile.

We tested whether secretion of apolipoprotein (apo) A-IV depends upon intestinal triglyceride (TG) transport by comparing output kinetics of TG and apo A-IV during and after duodenal lipid infusion in lymph-fistula rats. Lipid infusion (triolein, 40 mumol/h, 8 h) produced increases in lymphatic TG and apo A-IV output. After 8 h, triolein infusate was replaced with glucose-saline; TG output returned to basal levels 4-5 h later. However, apo A-IV output continued at significantly elevated levels until 20 h after the start of the experiment. Bile diversion blocked this continued output of A-IV during the post-lipid period, and resulted in basal TG output that was 75% lower than in bile-intact rats. Return of bile or low-dose triolein infusion (5 mumol/h) into the intestine reversed these effects. There were no differences in hepatic synthesis or filtration of plasma A-IV into lymph between bile-intact and bile-diverted groups. Intestinal A-IV synthesis was elevated in both groups even during the post-lipid period. The results support the hypothesis that intestinal triglyceride transport drives apo A-IV secretion, and suggest the existence of a bile-dependent, post-translational mechanism for the control of lymphatic apo A-IV output.

Animals↗

Large motor neuron involvement in Stiff-man syndrome: a qualitative and quantitative study.

Stiff-man syndrome (SMS) is characterized by fluctuating muscular rigidity and spasm. Recently, antibodies against glutamic acid decarboxylase (GAD), the enzyme catalyzing the synthesis of y-amino butyric acid (GABA), have been detected in SMS patients. An autoimmune mechanism against GAD was thus proposed for the suppression of GABAergic inhibitory interneurons, resulting in rigidity and spasm. We conducted quantitative investigations on the ventral horn of the spinal cord and its GAD immunoreactivity, post mortem, in a SMS patient and four controls. In the spinal cord of the SMS patient, we found a 70%, 33% and 27% reduction (P < 0.05) in the density of neurons with somal areas of 1000-1500 microm2, 500-1000 microm2, and 0-500 microm2, respectively. The density of neurons with a somal area greater than 1500 microm2 was not reduced, although some neurons in this class showed central chromatolytic changes. The affected muscles exhibited neurogenic atrophy. GAD-like immunoreactivity in the spinal gray matter was not significantly decreased. The density of Purkinje cells, known to contain high amounts of GAD, was not significantly reduced. While the co-occurrence of elevation of anti-GAD antibody in the serum and reduction in the density of small spinal neurons was confirmed, that of smaller alpha-motor neurons and gamma-motor neurons, the qualitative changes in larger alpha-motor neurons, and the preservation of spinal GAD-like immunoreactivity and non-spinal GAD-containing neurons suggest the involvement of factors other than autoimmune mechanisms through anti-GAD antibodies. More diverse mechanisms may be associated in the pathogenesis of SMS.

Cell Count↗