Expression of thymosin β4 in odontoblasts during mouse tooth development

Thymosin β4 (Tβ4) is expressed in developing tissue, where it stimulates cell differentiation and migration. Further, Tβ4 is expressed during molar development in mice, but the expression and function of Tβ4 in odontoblasts during mammalian tooth development have not yet been reported. Therefore, this study examined the expression and function of Tβ4 in differentiating odontoblasts during tooth development. As observed by immunohistochemistry, Tβ4 was expressed in the oral epithelium and inside cells of the tooth bud on embryonic day 15 (E15). Further, on E17, Tβ4 was expressed strongly in the dental lamina and oral epithelium, but only expressed in part of the cells in the outer and inner dental epithelium. Tβ4 was strongly expressed in the entire cytoplasm of odontoblasts on postnatal day 1 (PN1) and expressed intensively in the apical area of odontoblasts on PN4. Further, expression of Tβ4 was increased gradually in odontoblasts from PN1 to PN21. In an odontoblast cell line, MDPC-23, expression of Tβ4 mRNA and protein was increased strongly on day 4 and gradually decreased from day 14. The gene expression of dentin sialophosphoprotein (DSPP), bone sialoprotein (BSP), osteocalcin (OCN), osteonectin (ON), and collagen type I, related with mineralization, was significantly decreased in si-Tβ4/MDPC-23 during differentiation compared to that in MDPC-23 cells. Taken together, our results suggest that Tβ4 may be involved in oral epithelial cell proliferation at the initial stage of tooth development and regulates the expression and secretion of proteins during odontoblast differentiation.     

Association of variants in genes involved in pancreatic β-cell development and function with type 2 diabetes in North Indians

Variants in genes involved in pancreatic β-cell development and function are known to cause monogenic forms of type 2 diabetes and are also associated with complex form. In this study, we studied the genetic association of polymorphisms in such important genes with type 2 diabetes in the high-risk Indians. We genotyped 91 polymorphisms in 19 genes (ABCC8, HNF1A, HNF1B, HNF4A, INS, INSM1, ISL1, KCNJ11, MAFA, MNX1, NEUROD1, NEUROG3, NKX2.2, NKX6.1, PAX4, PAX6, PDX1, USF1 and WFS1) in 2025 unrelated North Indians of Indo-European ethnicity comprising of 1019 diabetic and 1006 non-diabetic subjects. HNF4A promoter P2 polymorphisms rs1884613 and rs2144908, which are in high linkage disequilibrium, showed significant association with type 2 diabetes (odds ratio (OR)=1.37 (95% confidence interval (CI) 1.19-1.57), P=9.4 × 10(-6) for rs1884613 and OR=1.37 (95%CI 1.20-1.57), P=6.0 × 10(-6) for rs2144908), as previously shown in other populations. We observed body mass index-dependent association of these variants with type 2 diabetes in normal-weight/lean subjects. Variants in USF1, ABCC8, ISL1 and KCNJ11 showed nominal association, while haplotypes in these genes were significantly associated. rs3812704 upstream of NEUROG3 significantly increased risk for type 2 diabetes in normal-weight/lean subjects (OR=1.68 (95%CI 1.25-2.24), P=4.9 × 10(-4)). Thus, pancreatic β-cell development and function genes contribute to susceptibility to type 2 diabetes in North Indians.     

NF-κB in control of regulatory T cell development,identity, and function

Journal of Molecular Medicine - Regulatory T cells (Treg cells) act as a major rheostat regulating the strength of immune responses, enabling tolerance of harmless foreign antigens, and preventing...     

Postnatal expression of transport proteins involved in acid–base transport in mouse kidney

The kidney plays a major role in maintaining and controlling systemic acid–base homeostasis by reabsorbing bicarbonate and secreting protons and acid-equivalents, respectively. During postnatal kidney development and adaptation to changing diets, plasma bicarbonate levels are increasing, the capacity for urinary acidification maturates, and the final morphology and distribution of intercalated cells is achieved. In adult kidney, at least two types of intercalated cells (IC) are found along the collecting duct characterised either by the expression of AE1 (type A IC) or pendrin (non-type A IC) where non-type A IC are found only in the convoluted distal tubule, connecting tubule and cortical collecting duct. Here we investigated in mouse kidney the relative mRNA abundance, protein expression levels and distribution of several proteins involved in renal acid–base transport, namely, the Na⁺/HCO₃ ^– cotransporter NBC1 (SLC4A4), the Na⁺/H⁺-exchanger NHE3 (SLC9A3), two subunits of the vacuolar H⁺-ATPase [ATP6V0A4 (a4), ATP6V1B1 (B1)], the Cl^–/HCO₃ ^– exchangers AE1 (SLC4A1) and pendrin (SLC26A4). Relative mRNA abundance of all transport proteins was lowest at day 3 after birth and increased thereafter in parallel with protein levels. The numbers of type A and non-type A IC in the cortical collecting duct (CCD) increased from day 3 to days 18 and 24, whereas the number of IC in the CCD with apical staining for the vacuolar H⁺-ATPase subunits a4 and B1 decreased from day 3 to days 18 and 24, respectively. In addition, cells with characteristics of non-type A IC (pendrin expression, basolateral expression of vacuolar H⁺-ATPase subunits) were found in the inner and outer medulla 3 days after birth but were absent from the medulla of 24-day-old mice. Taken together, these results demonstrate massive changes in mRNA and protein expression levels of several acid–base transporters during postnatal kidney maturation and also show changes in intercalated cell phenotype in the medulla during these processes.     

Expression of FGF10 and FGFR2Ⅲb in development of mouse kindey

目的 观察FGF10及其受体FGFR2Ⅲb在小鼠肾脏发生发育中的表达和分布,探讨其在肾脏发生发育中的作用.方法 选取胚龄12、14、16及18 d和出生后1、7、14、21及42 d小鼠肾组织,用免疫组织化学和Western blot,检测FGF10和FGFR2Ⅲb的表达.结果 胚龄12 d组FGF10及FGFR2Ⅲb开始在输尿管芽微弱表达;随着肾脏发育成熟,FGF10及FGFR2Ⅲb主要表达于远端小管和集合管;FGF10在近曲小管表达,近直小管无表达;FGFR 2Ⅲb在近端小管无阳性表达;生后肾组织及发育各阶段肾小体未见FGF10及FGFR2Ⅲb表达.Western blot显示随着胚(日)龄的增加,FGF10及FGFR2Ⅲb在肾脏的表达量先增后减.结论 FGF10和FGFR2Ⅲb在肾脏发生发育过程中发挥重要作用.     

α- and β-Adducin polymorphisms affect podocyte proteins and proteinuria in rodents and decline of renal function in human IgA nephropathy

Adducins are cytoskeletal actin-binding proteins (α, β, γ) that function as heterodimers and heterotetramers and are encoded by distinct genes. Experimental and clinical evidence implicates α- and β-adducin variants in hypertension and renal dysfunction. Here, we have addressed the role of α- and β-adducin on glomerular function and disease using β-adducin null mice, congenic substrains for α- and β-adducin from the Milan hypertensive (MHS) and Milan normotensive (MNS) rats and patients with IgA nephropathy. Targeted deletion of β-adducin in mice reduced urinary protein excretion, preceded by an increase of podocyte protein expression (phospho-nephrin, synaptopodin, α-actinin, ZO-1, Fyn). The introgression of polymorphic MHS β-adducin locus into MNS (Add2, 529R) rats was associated with an early reduction of podocyte protein expression (nephrin, synaptopodin, α-actinin, ZO-1, podocin, Fyn), followed by severe glomerular and interstitial lesions and increased urinary protein excretion. These alterations were markedly attenuated when the polymorphic MHS α-adducin locus was also present (Add1, 316Y). In patients with IgA nephropathy, the rate of decline of renal function over time was associated to polymorphic β-adducin (ADD2, 1797T, rs4984) with a significant interaction with α-adducin (ADD1, 460W, rs4961). These findings suggest that adducin genetic variants participate in the development of glomerular lesions by modulating the expression of specific podocyte proteins.     

Changes of calcium binding proteins,c-Fos and COX in hippocampal formation and cerebellum of Niemann–Pick,type C mouse

Niemann–Pick disease, type C (NPC) is an intractable disease that is accompanied by ataxia, dystonia, neurodegeneration, and dementia due to an NPC gene defect. Disruption of calcium homeostasis in neurons is important in patients with NPC. Thus, we used immunohistochemistry to assess the expression levels of calcium binding proteins (calbindin D28K, parvalbumin, and calretinin), c-Fos and cyclooxygenase-1,2 (COX-1,2) in the hippocampal formation and cerebellum of 4 and 8 week old NPC+/+, NPC+/−, and NPC−/− mice.General expression of these proteins decreased in the hippocampus and cerebellum of NPC−/− compared to that in both young and adult NPC+/+ or NPC+/− mice. Parvalbumin, COX-1,2 or c-Fos-immunoreactive neurons were widely detected in the CA1, CA3, and DG of the hippocampus, but the immunoreactivities were decreased sharply in all areas of hippocampus of NPC−/− compared to NPC+/+ and NPC+/− mice.Taken together, reduction of these proteins may be one of the strong phenotypes related to the neuronal degeneration in NPC−/− mice.     

Immunohistochemical analysis of various serum proteins in living mouse thymus with “in vivo cryotechnique”

It has been difficult to clarify the precise localizations of soluble serum proteins in thymic tissues of living animals with conventional immersion- or perfusion-fixation followed by alcohol dehydration owing to ischemia and anoxia. In this study, ??in vivo cryotechnique?? (IVCT) followed by freeze-substitution fixation was performed to examine the thymic structures of living mice and immunolocalizations of intrinsic or extrinsic serum proteins, which were albumin, immunoglobulin G1 (IgG1), IgA, and IgM, as well as intravenously injected bovine serum albumin (BSA). Mouse albumin was more clearly immunolocalized in blood vessels and interstitial matrices of the thymic cortex than in tissues prepared by the conventional methods. The immunoreactivities of albumin and IgG1 were stronger than those of IgA and IgM in the interstitium of subcapsular cortex. The injected BSA was time-dependently immunolocalized in blood vessels and the interstitium of corticomedullary areas at 3.5 h after its injection, and then gradually diffused into the interstitium of the whole cortex at 6 h and 12 h. Thus, IVCT revealed definite immunolocalizations of serum albumin and IgG1 in the interstitium of thymus of living mice, indicating different accessibility of serum proteins from the corticomedullary areas, not from the subcapsular cortex of living animals, depending on various molecular sizes and concentrations.     

Effects of internal chloride on ATP-sensitive K-channels in mouse pancreatic β-cells

The effect of internal Cl^– on the K-ATP channel of pancreatic ß-cells was examined. Reducing Cl^– from 140mM to 14mM potentiated channel activity (NP_o) 4.5 fold in 60% of patches but was without effect in 40% of patches. The K_i for channel inhibition by ATP was not changed. The inhibitory effect of Cl^– interacted with the stimulatory action of MgGDP. In 140mM Cl^–, ImM MgGDP increased NP_o in Cl^–-sensitive patches only: when NP_o was potentiated by 14mM Cl^–, no further increase was produced by MgGDP. These observations suggest that MgGDP and low Cl^– solution may increase channel activity via a similar mechanism.     

Role of CD3ε-mediated signaling in T-cell development and function

Receptor-mediated signal transduction plays an important role in T-cell differentiation and function. The pre-T-cell receptor (pre-TCR) and TCR complexes are the most critical receptors for T-cell biology. Signals induced by pre-TCR and TCR in a ligand-independent or a ligand-dependent manner, respectively, are essential for thymocyte maturation. CD3 proteins, which are γ, δ, ε, and ζ polypeptides, play pivotal role in intracellular assembly, surface expression, and signal transduction via the pre-TCR and TCR complexes. Recent studies have suggested central and multiple roles for CD3ε in T-cell development and function. We review the role of the CD3ε chain in T-cell biology.     

Reduction of β-amyloid pathology by celastrol in a transgenic mouse model of Alzheimer's disease

Background  

Aβ deposits represent a neuropathological hallmark of Alzheimer's disease (AD). Both soluble and insoluble Aβ species are considered to be responsible for initiating the pathological cascade that eventually leads to AD. Therefore, the identification of therapeutic approaches that can lower Aβ production or accumulation remains a priority. NFκB has been shown to regulate BACE-1 expression level, the rate limiting enzyme responsible for the production of Aβ. We therefore explored whether the known NFκB inhibitor celastrol could represent a suitable compound for decreasing Aβ production and accumulation in vivo.     

Up-regulation of novel intermediate filament proteins in primary fiber cells: an indicator of all vertebrate lens fiber differentiation?

The early embryonic development and expression patterns of the eye lens specific cytoskeletal proteins, CP49 and CP95, were determined for the chick and were found to be similar in both human and mouse. These proteins, as well as their homologs in other species, are obligate polymerization partners which form unique filamentous structures termed "beaded filaments." CP49 and CP95 appeared as protein products after 3 days of embryonic development in the chick during the elongation of primary fiber cells. Although limited data were obtained for human embryos at these early developmental timepoints, they were consistent with the interpretation that the up-regulation of these lens specific proteins began only after the initiation of lens vesicle closure. In situ hybridization with the mouse lens confirmed that message levels for beaded filament proteins were greatly elevated in differentiating primary fiber cells. Nuclease protection assays established that mRNA levels for CP49 remained relatively constant while CP95 mRNA levels increased once the process of secondary fiber formation was under way. Although present in relatively low abundance, the mRNA for a unique splice variant of CP49, CP49(INS), was also detected early in embryonic development and into adulthood. Peptide-specific antibodies directed against unique predicted sequences were able to confirm the protein expression of CP49(INS) in both embryonic and adult chick lens cells. These data present the first detailed study of the expression of CP49 and CP95 during early lens development. They suggest that the up-regulated expression of CP49 and CP95 could serve as pan-specific markers for all vertebrate lens fiber development.     

Increase in IL-6, TNF-α, and MMP-9, but not sICAM-1, concentrations depends on exercise duration

It has been suggested that exercise intensity is of importance in the regulation of increase in pro-inflammatory molecules, but there is still a debate about the effect of duration on these molecules. Therefore, the effect of exercise duration on the serum concentrations of interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α), soluble form of intercellular adhesion molecule-1 (sICAM-1), and matrix metalloproteinase-9 (MMP-9) was studied in 22 half-marathon (HM) and 18 marathon (M) male amateur runners who completed their exercise task in 1.8 ± 0.2 (mean ± standard deviation) and 3.6 ± 0.4 h, respectively (thus, average speed was 11.7 ± 1.5 and 11.9 ± 1.8 km h^?1, respectively). Blood was sampled 2 days before, 15 min after, and 28 h after the race. IL-6, TNF-α, and MMP-9 always increased immediately after exercise, but the increase was larger (P < 0.05) in M versus HM (?IL-6: 31 ± 24 vs. 5 ± 4 pg ml^?1; ?TNF-α: 1.7 ± 1.9 vs. 0.5 ± 0.8 pg ml^?1; MMP-9: 288 ± 216 vs. 145 ± 128 ng ml^?1, respectively). sICAM-1 also increased with exercise, but similarly in M and HM (20 ± 40 vs. 23 ± 32 ng ml^?1, respectively). Only sICAM-1 remained elevated 28 h post-exercise in both HM and M, while IL-6, TNF-α, and MMP-9 returned to pre-exercise levels. Competitive HM and M races induce significant increases in IL-6, TNF-α, sICAM-1, and MMP-9 concentrations. As HM and M runners performed the competition with similar absolute intensity, the difference in response between the groups suggests that exercise duration is of importance in the regulation of IL-6, TNF-α, and MMP-9, but not sICAM-1 concentrations in response to prolonged running.     

Adeno-associated virus-mediated expression of β-hexosaminidase prevents neuronal loss in the Sandhoff mouse brain

Sandhoff disease, a GM2 gangliosidosis caused by a deficiency in β-hexosaminidase, is characterized by progressive neurodegeneration. Although loss of neurons in association with lysosomal storage of glycosphingolipids occurs in patients with this disease, the molecular pathways that lead to the accompanying neurological defects are unclear. Using an authentic murine model of GM2 gangliosidosis, we examined the pattern of neuronal loss in the central nervous system and investigated the effects of gene transfer using recombinant adeno-associated viral vectors expressing β-hexosaminidase subunits (rAAV2/1-Hex). In 4-month-old Sandhoff mice with neurological deficits, cells staining positively for the apoptotic signature in the TUNEL reaction were found in the ventroposterior medial and ventroposterior lateral (VPM/VPL) nuclei of the thalamus. There was progressive loss of neuronal density in this region with age. Comparable loss of neuronal density was identified in the lateral vestibular nucleus of the brainstem and a small but statistically significant loss was present in the ventral spinal cord. Loss of neurons was not detected in other regions that were analysed. Administration of rAAV2/1-Hex into the brain of Sandhoff mice prevented the decline in neuronal density in the VPM/VPL. Preservation of neurons in the VPM/VPL was variable at the humane endpoint in treated animals, but correlated directly with increased lifespan. Loss of neurons was localized to only a few regions in the Sandhoff brain and was prevented by rAAV-mediated transfer of β-hexosaminidase gene function at considerable distances from the site of vector administration.     

Upregulation of β-catenin expression in down syndrome model Ts65Dn mouse brain

Trisomy of human chromosome 21 (Hsa 21) causes the pathological characteristics of Down syndrome (DS). Little is known about the mechanisms by which trisomy 21 affects the expression of genes on other chromosomes. Using a mouse model of DS, the Ts65Dn mouse, we have performed mRNA and protein measurements to identify genes on chromosomes not syntenic with Hsa 21 whose expression is affected by the presence of three copies of genes between loci Mrpl39 and Znf295 on mouse chromosome 16 (Mmu 16). We report the upregulation of β-catenin, located on mouse chromosome 9 (Mmu 9) in Ts65Dn brain. Using immunocytochemistry on Ts65Dn and control mouse brain tissue, we observed a striking increase in β-catenin expression specifically in the endothelial cells lining the cerebral blood vessels of the Ts65Dn mice. Since β-catenin is involved in cell–cell adhesion, upregulation of this protein in DS may alter adherens protein interactions that are involved in the normal functions of endothelial cells. Elevated β-catenin might be responsible for altered endothelial cell function/s leading to the impairment of brachial flow velocity observed in DS.     

TNFα-dependent development of lymphoid tissue in the absence of RORγt+ lymphoid tissue inducer cells

Lymphoid tissue often forms within sites of chronic inflammation. Here we report that expression of the proinflammatory cytokine tumor necrosis factor α (TNFα) drives development of lymphoid tissue in the intestine. Formation of this ectopic lymphoid tissue was not dependent on the presence of canonical RORgt⁺ lymphoid tissue–inducer (LTi) cells, because animals expressing increased levels of TNFα but lacking RORgt⁺ LTi cells (TNF/Rorc(gt)^−/− mice) developed lymphoid tissue in inflamed areas. Unexpectedly, such animals developed several lymph nodes (LNs) that were structurally and functionally similar to those of wild-type animals. TNFα production by F4/80⁺ myeloid cells present within the anlagen was important for the activation of stromal cells during the late stages of embryogenesis and for the activation of an organogenic program that allowed the development of LNs. Our results show that lymphoid tissue organogenesis can occur in the absence of LTi cells and suggest that interactions between TNFα-expressing myeloid cells and stromal cells have an important role in secondary lymphoid organ formation.