MicroRNA-15a/β1,4-GalT-I axis contributes to cartilage degeneration via NF-κB signaling in osteoarthritis

Abstract Objective Osteoarthritis is a condition characterized by articular cartilage degradation. The increased expression of β1,4-Galactosyltransferase-I (β1,4-GalT-I) in the articular cartilage of osteoarthritis patients was related to an inflammatory response. The aim of this study was to elucidate the role of β1,4-GalT-I in osteoarthritis. This study aimed to determine the function of 1,4-GalT-I in osteoarthritis. Methods The osteoarthritis mouse model with the destabilization of the medial meniscus was established by microsurgical technique. Pathological changes in articular cartilage were observed by hematoxylin and eosin staining and safranin O-fast green staining. Quantitative real-time polymerase chain reaction, western blot, and enzyme-linked immunosorbent assays were used to observe mRNA and protein expression, respectively. RNA interactions were verified by a luciferase reporter assay. SA-β-Gal staining was used to assess chondrocyte senescence. Immunofluorescence staining was conducted to observe the localization of Nuclear Factor-kappaB (NF-κB). Results β1,4-GalT-I and microRNA-15a (miR-15a) show high and low expression in the articular cartilage of osteoarthritis, respectively. MiR-15a inhibits the mRNA translation of β1,4-GalT-I. β1,4-GalT-I promotes extracellular matrix degradation, senescence, and NF-κB activation in IL-1β-stimulated chondrocytes, which can be reversed by overexpression of miR-15a. Intra-articular injection of microRNA-15a ameliorates cartilage degeneration by inhibiting β1,4-GalT-I and phosphorylation of NF-κB in vivo. Conclusion The authors clarified that the miR-15a/β1,4-GalT-I axis inhibits the phosphorylation of NF-κB thereby inhibiting extracellular matrix degradation and senescence in chondrocytes to alleviate cartilage degeneration in osteoarthritis. MiR-15a and β1,4-GalT-I may serve as potentially effective targets for the future treatment of osteoarthritis.

Anti-PP₁P(k) (Tj(a)) Antibody in a Korean Female Patient with p Phenotype Confirmed by Genotyping

Clinical, biochemical and molecular profile of variant Galactosemia in children detected by National Newborn Screening: A pilot study

Objective@#The observed irregularities in the biochemical profile and the limited information on long-term outcomes among patients with Duarte variant (D/G) galactosemia have led to patient management variability. This study examined the molecular characteristics of Filipino patients with presumed variant galactosemia for confirmation of diagnosis. It also aimed to describe the corresponding biochemical, clinical and neurodevelopmental profiles in order to gain a better understanding of the patients with normal galactose metabolites in spite of low to absent GALT activity detected by the local newborn screening program. @*Methods@#Thirteen (13) patients who were presumed to have a variant form of galactosemia by national newborn screening between 2002 and 2010, and who previously underwent physical and neurodevelopmental assessment were included in the study. Repeat clinical, ophthalmologic and neurodevelopmental evaluations were done upon recruitment of participants. Direct sequence analysis of the coding region of the GALT gene was conducted to determine the patients’ genotypes. @*Results@#None of the patients’ genotypes were consistent with Duarte variant (D/G) galactosemia. Their genotypes reflect the normal total blood galactose levels in patients, but were inconsistent with the absent or trace GALT activity.@*Conclusion@#Molecular testing for the entire cohort of presumed “variant” galactosemia Filipino patients will provide better profiling of this condition. Re-evaluation and assessment of the current guidelines used by national newborn screening in classifying variant galactosemia are recommended.

Association of C1GALT1 gene polymorphisms with susceptibility and prognosis of IgA nephropathy on Uyghur population in Xinjiang Uyghur Autonomous Region / 中华肾脏病杂志

Objective To explore the genetic association of C1GALT1 gene polymorphisms with susceptibility and prognosis of IgA nephropathy (IgAN) on Uyghur population in Xinjiang Uyghur Autonomous Region.Methods Ninety Uighur patients with IgAN and ninety geographically and age matched healthy controls were recruited.Peripheral blood was collected from recruited individuals for DNA extracting.After amplified by polymerase chain reaction (PCR),genotyping of the four single nucleotide polymorphisms (SNPs) in C1GALT1 gene,which were rs9639031,rs5882115,rs1008898,-527A/G,were detected by direct sequencing analysis.Differences of allele and genotype frequency were analyzed between IgAN and healthy controls.Moreover,the association between these SNPs and the risk and progress in IgAN patients were further analyzed.Results (1) The I allele frequency of rs5882115 in C1GALT1 gene was significantly higher in IgAN than that in healthy controls （x2 =7.788,P =0.015),no difference in allele frequencies of rs9639031,rs1008898,-527A/G between IgAN and healthy controls was found.Under the dominant mode,the DI+ Ⅱ genotype frequencies of rs5882115 was significantly higher in IgAN than that in healthy controls (x2 =8.563,P =0.009),no difference in genotype frequencies of rs9639031,rs1008898,-527A/G between IgAN and healthy controls was found.Under the hidden mode,no difference in genotype frequencies of rs5882115,rs9639031,rs1008898,-527A/G between IgAN and healthy controls was found.Logistic single factor regression analysis showed that the risk to IgAN of whom carry I allele of rs5882115 was 2.469 times than the D allele (OR =2.469),and the risk to IgAN of whom carry DI+ Ⅱ genotype was also higher (OR =2.852).(3) No association was found between these SNPs in C1GALT1 gene and serum creatinine in IgAN.Conclusion Association between rs5882115 in C1GALT1 gene and Uighur IgAN susceptibility suggests that there may be variants in C1GALT1 gene or its linked genetic region,which needs further exploration.

Participación de las células Th17 en la patogenia de la infección por el virus de la inmunodeficiencia humana de tipo 1 / Th17 cells involvement in the pathogenesis of the human immunodeficiency virus type 1

La infección por el VIH-1 se caracteriza por la eliminación de linfocitos T CD4+, particularmente en la mucosa gastrointestinal, que favorece la traslocación microbiana y la hiperactivación inmunitaria, principal mecanismo patogénico en esta infección. Las células Th17 son una subpoblación proinflamatoria de linfocitos CD4+, que producen IL-17, IL-21 e IL-22, y son importantes en la respuesta antimicrobiana, principalmente en el sistema gastrointestinal, donde promueven la restauración de la mucosa. Aunque su eliminación se ha asociado con progresión de la infección por el VIH-1 y por el virus de la inmunodeficiencia de los simios, y han sido descritas como deletérea en autoinmunidad. Su papel en la patogenia de la infección por el VIH-1 no está claramente establecido. Considerando su capacidad funcional, las células Th17 podrían tener un impacto dual, dependiendo de la fase de la infección en que se encuentre el individuo. Actualmente, hay más información que sugiere que estas células tienen un papel benéfico al promover la recuperación de la mucosa intestinal y disminuir la traslocación microbiana, así como la hiperactivación inmunitaria. Sin embargo, su papel patogénico, particularmente promoviendo la replicación viral mediante la producción de citocinas proinflamatorias, no debe descartarse. En esta revisión, se presentan los datos científicos disponibles del efecto de las células Th17 en la patogenia de la infección por el VIH-1.

HIV-1 infection is characterized by a gradual decrease of the immunological competence and a massive depletion of CD4+ T cells, particularly in gut-associated lymphoid tissue, which leads to microbial translocation, contributing to immune hyperactivation, the main pathogenic mechanism during HIV-1 infection. Th17 cells are a proinflammatory CD4+ T cell subset, which produce IL-17, IL-21 and IL-22 and play a pivotal role in host defense, mainly in the gastrointestinal tissue, where they promote antimicrobial responses and gut mucosa restoration. Although Th17 depletion is a hallmark of the progression of the simian and human immunodeficiency viral infections and they have been involved in the pathogenic process in some autoimmune diseases, the role of these cells during HIV-1 infection is not completely understood. Considering their functional potential, Th17 cells could have a dual role, depending on the stage of HIV infection a patient has reached. Currently, most evidence suggests that Th17 cells have a beneficial role by promoting gut mucosa recovery, preventing microbial translocation and decreasing immune hyperactivation. However, the pathogenic role of these cells, particularly, increasing viral replication through the production of inflammatory cytokines should not be ruled out. In this review, scientific evidence regarding the role of Th17 on the pathogenesis of HIV infection is discussed.

Microbes in the gut A digestable account of host-symbiont interactions.

The human bowel is host to a diverse group of bacteria with over 500 different bacterial species contributing to this diversity. Until recently these bacteria were regarded as residents without any specific functions. The last two decades have seen a radical change in our understanding of the interactions between the gut flora and their eukaryotic hosts and there is a growing appreciation of the spectrum of functions performed by these symbionts. Intestinal bacteria are recognized for their role in nutrient absorption, mucosal barrier function, angiogenesis, morphogenesis and postnatal maturation of intestinal cell lineages, intestinal motility and more importantly maturation of gut associated lymphoid tissue (GALT). Although gut flora are implicated in certain pathological disorders, their remarkable contributions to health and homeostasis of the host need to be recognized and understood.

A Case of Classical Galactosemia caused by Compound Heterozygous Mutations of the GALT Gene

Classical galactosemia is an autosomal recessive disorder of galactose metabolism, caused by a deficiency of the enzyme galactose-1-phosphate uridyltransferase (GALT). Buildup of galactose-1-phosphate is toxic at high levels and can damage the liver, brain, eyes, and other vital organs. The case presented here was that of an 11-day-old female infant who had elevated galatose levels upon initial neonatal screening test with persistent cholestatic jaundice, coagulopathy, and hepatomegaly. The patient was transferred due to aggravation of clinical symptoms including bleeding and jaundice. She had a delayed galactose free diet because of an inappropriate diagnosis. We quickly provided her with a lactose/ galactose-restricted diet as per her final diagnosis. Clinical and laboratory results were improved after a few days of treatment. For confirmatory testing for classical galactosaemia, we simultaneously analyzed for GALT enzyme activity and allele-specific PCR/fragments for seven mutations and two polymorphisms in the GALT gene. We were able to find several GALT-deficient and compound heterozygous mutations of the GALT gene.

Development of apoptosis after small bowel transplantation in rats / 中国康复理论与实践

@#ObjectiveTo investigate the development of apoptosis during ischemia/reperfusion (IR) injury and acute rejection, and to explore the significance of apoptosis in the Graft Mesenteric Lymph Node (GALT) in a rat heterotopic small bowel transplant (SBT) model.MethodsSBT was performed in F344/N rats with either freshly harvested or preserved (4 h, in ringer lactate solution at 4 ℃) syngeneic and allogeneic (Wistar/A-F344/N) grafts. Bowel and GALT samples were collected 2 h after reperfusion and on small bowel transplant postoperative days (POD) 1, 4, and 7. Histopathology assessment of the graft and GALT were prepared for hematoxylin-eosin (H&E) staining. Apoptosis was detected by the TUNEL and the electron microscope. ResultsThe number of apoptotic cells 2 h after reperfusion increased profoundly in association with preservation. After a significant decrease on POD 1, the apoptotic cells rose again between POD 3 and 7 only in allogeneic grafts. On the other hand, the apoptotic cells in allogeneic GLAT markedly increased from POD 1 to day 3; at that time point, neither histological findings of rejection nor increase in apoptotic crypt cells were present in the graft jejunum. ConclusionIR injury and acute rejection may both induce extensive apoptosis. The graft jejunum distinct second increase in apoptosis may be an early and specific sign of acute rejection. Apoptosis of GLAT cells was well correlated with and ahead of progression of acute rejection.

Galactosemia Detected by Neonatal Screening Test

PURPOSE: The genetic disturbance of galactosemia is expressed as a cellular deficiency of either galactose-1-phosphate uridyltransferase(GALT) or galactokinase(GALK) or UDP galactose 4-epimerase(GALE). To find-out the pattern of galactosemia in Korea, we retrospectively analyzed cases of galactosemia detected by neonatal screening program. METHODS: We analyzed medical records of patients who visited Soonchunhyang University Hospital at age of 1 month after showing abnormalities in neonatal screening of galactosemia. For accurate diagnosis, galactose was measured by enzyme immunoassay(EIA) and fluorophotometer, also galactose-1-phosphate by fluorophotometer. Enzyme activities of GALK, GALT and GALE in RBC and galactose-1-phosphate were measured by radioisotope assay(RIA). Beutler test were done. Patients went on a lactose-free diet and follow-up tests for galactose, galactose-1-phosphate level and enzyme activity were performed. RESULTS: 10 patients(male : 6, female : 4) were diagnosed as galactosemia. Two patients had GALK deficiency and two had GALT deficiency. Six were GALE deficient showing the largest number. In two patients with GALK deficiency, GALT and GALE activities were normal but GALK activities showed respectively reduced activity. For GALT deficiency, two patients had low GALT activity in RBC and showed genotype of Duarte 2/G(galactosemia) in DNA analysis. In one patient, GALT activity was normal. Three patients seemed to be heterozygote state of GALE deficiency according to GALE activity levels. Four patients showed GALK hyperactivity. CONCLUSION: GALE deficiency provided the highest number. After lactose-free diet, galactose and galactose-1-phosphate were normaly maintained. Neonatal screening on galactosemia is essential for preventing life-threatening symptoms and an accurate diagnosis is needed for finding out the type of galactosemia which is important for prognosis.

Factors Related to Increased CA19-9 andLewis Antigen in Pancreatic Cancer Cell Lines

PURPOSE: The 8 pancreatic cancer cell lines (BxPC-3, Capan-2, CFPAC-1, HPAC, Capan-1, AsPC-1, MIA PaCa-2, and PANC-1) were investigated to identify the factors which would increase CA19-9 related to the Lewis antigen. CA19-9 in serum is a well-known tumor marker, and is frequently used for the clinical diagnosis of pancreatic cancer. The oligosaccharide on the CA19-9 epitope is a sialylated Lewis A blood group antigen. METHODS: beta3Gal-T was detected by reverse transcriptase polymerase chain reaction (RT-PCR). The phenotypes and genotypes of Lewis antigen were determined by flow cytometry analysis and restriction fragment length polymorphism (RFLP), respectively. The phenotypes of sLe(a) were assessed by flow cytometry analysis and the sLe(a) on supernatants was detected by sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE). CA19-9 and DUPAN-2 on supernatants were measured by enzyme immunoassay. RESULTS: CA19-9 productions were possible from all cell lines since they all had beta3Gal-T and there were no genotypical Lewis negative (le/le). The elevation of CA19-9 was noted on Capan-2 and CFPAC-1, which were phenotypically Lewis positive (Le(a+b+)), as expected. Interestingly, it was also elevated in BxPC-3 even though the line was known to be phenotypically Lewis negative (Le(a-b-)). Sialyl Le(a) appeared to play an important role in this phenomenon. Although CA 19-9 was not detected in the phenotypically Lewis negative pancreatic cell line without sialyl Le(a), the levels of DUPAN-2 were variable. CONCLUSION: It was revealed that an elevated CA19-9 was related with increased expression of Lewis gene, not merely the existence of the gene. Further investigations on the role of ST3Gal are warranted to explain the mechanisms of the variable levels of DUPAN-2 in Le(a-b-) cell lines.

Detection of ASC in spleen cells and GALT after immunization with two bivalent Shigella vaccines in different administration rout in mice / 中国免疫学杂志

Abstract Objective:FSM-2117 and FS-5416 are two bivalent hybrid strains of S.flexneri and S.sonnei, constructed by this laboratory-The FS-5416 expresses four invassive plasmid antigens(IpaA、IpaB、IpaC and IpaD) and has a good contact heamolysis activity(CHA+ ), butFSM-2117 hasn' t. To observe the immunogenicity of Shigelk vaccines through three different mucosal administration rout, the changes of ASCin the spleen and Peyer's patch(PP) , mesenteric lymph nodes(MLN) lymphocytes are detected.Methods:BALB/c mice were divided intothree groups, 20 mice per group. Mice were immunized respectively with the Ipa+ or Ipa- vaccines(4 x 10~7 CFU) three times with an intervalof two weeks by intranasal、intragastric or intraintestinal administration routs. The spleen , PP , MLN lymphocytes were isolated of seventh dayat random after immunization. An BA-EIISASPOT was done to account the numbers of ASC.Results:The numbers of SIgA and SIgG ASC ofspleen , PP , MLN lymphocytes of intranasal and intraintestinal group were significantly increased . Significant difference were only observed inthe number of spleen lymphocytes SIgA and SIgG ASC of intranasal group between Ipa+ and Ipa- . Conclusion:Two bivalent Shigelk vaccinescan induce spleen and GALT immunity reaction by nasal or small intestinal mucosal in a low dosage compared with intragastric rout (about 1/20). Ipa can significantly increased the number of spleen lymphocytes SIgA and SIgG ASC of intranasal group.

Biochemical analysis of galactose induced bacteriostasis in galT mutants of Escherichia coli K 12.

Escherichia coli mutants completely defective in galactose- 1-phosphate uridyl transferase (EC 2.7.7.10) and growing in glycerol medium undergo rapid cessation of growth when exposed to galactose. Toxicity due to galactose is equally pronounced when glycerol is replaced by other carbon sources, like succinate and proline. Gas chromatographic analysis failed to detect even trace amounts of galactitol. Moreover, galactose- 1-phosphate had no inhibitory role on some of the critical enzymes of cellular metabolism. General loss of energy (ATP) due to futile phosphorylation of galactose is probably the cause of bacteriostasis. The galT mutants can serve as models of human transferaseless galactosemia only to a limited extent.