Expression and cellular distribution of TLR4, MyD88, and NF-κB in diabetic renal tubulointerstitial fibrosis, in vitro and in vivo

Aim

Inflammation and extracellular matrix hyperplasia are crucial in the pathogenesis of tubulointerstitial fibrosis (TIF) involved in diabetic nephropathy (DN). Macrophage accumulation plays a major role, but whether immune factors contribute to DN pathogenesis is not well understood. This study aimed to investigate TLR4-MyD88-NF-κB-dependent pathway's involvement in TIF pathogenesis.

Methods

STZ-induced diabetic rats and rat renal tubular epithelial NRK-52E cells cultured under high glucose conditions were used as in vivo and in vitro models, respectively. Real-time RT-PCR, western blot, immunohistochemistry and immunofluorescence were performed to examine the mRNA and protein levels of TLR4, MyD88, NF-κB, MCP-1, and α-SMA.

Results

Compared with 5.5 mmol/L glucose, treatment of NRK-52E cells with 25 and 50 mmol/L d-glucose resulted in significantly increased TLR4 and MyD88 mRNA and protein levels (P < 0.05). TLR4 and MyD88 were detected in the cytoplasm of most NRK-52E cells cultured under high glucose. Pronounced damage in the renal tubulointerstitium was observed in diabetic rats (scores: 3.82 ± 0.65 vs. 0.38 ± 0.08, P < 0.01). Compared with the normal controls, a sharp upregulation of TLR4, MyD88, NF-κB p65, MCP-1, and α-SMA mRNA and protein levels was observed in diabetic rat kidneys (P < 0.05). In diabetic animals, TLR4 and MyD88 were strongly expressed in the cytoplasm, while NF-κB p65 was widely expressed in cytoplasm and nuclei of renal tubular epithelial cells.

Conclusion

The inflammatory reaction and epithelial-mesenchymal transformation observed in renal tubulointerstitium may be the result of overactivation of the TLR4-MyD88-NF-κB-dependent innate immunity under high glucose, and may be involved in DN occurrence and progression.     

比较踝关节骨折后踝螺钉不同进钉方向固定疗效

目的 比较踝关节骨折后侧入路后踝螺钉不同进钉方向的固定疗效。 方法 选取漂浮体位下后外侧入路对后踝骨折分别采用不同进钉方向螺钉固定患者48例,男性20例,女性28例,年龄（51.88 ± 7.90）岁,左踝20例,右踝28例,依据Lauge-Hansen踝关节骨折分型：旋后外旋型Ⅳ度36例,旋前外旋型Ⅳ度12例。后踝均采用螺钉固定,前向后固定组22例,后向前固定组26例,对比两组手术时间、骨折愈合时间及并发症情况,末次随访采用AOFAS评分进行临床疗效评价。 结果 两组病例均获随访,随访时间10~20个月,平均（13.0±3.5）个月,均Ⅰ期愈合,无切口感染、血管神经损伤等并发症,手术时间、骨折愈合时间、AOFAS评分均未见统计学差异,P>0.05。 结论 后外侧入路固定后踝骨折疗效较好,螺钉不同进钉方向具有相似的临床固定结果,手术医生可凭经验选择合适的方法。     

Zinc induces protein phosphatase 2A inactivation and tau hyperphosphorylation through Src dependent PP2A (tyrosine 307) phosphorylation

The activity of protein phosphatase (PP) 2A is downregulated and promotes the hyperphosphorylation of tau in the brains of Alzheimer's disease (AD), but the mechanism for PP2A inactivation has not been elucidated. We have reported that PP2A phosphorylation at tyrosine 307 (Y307) is involved in PP2A inactivation. Here, we further studied the upstream mechanisms for PP2A phosphorylation and inactivation. We found that zinc, a heavy metal ion that is widely distributed in the normal brain and accumulated in the susceptible regions of AD brain, could induce PP2A inhibition, phosphorylation of PP2A at Y307 and tau hyperphosphorylation both in rat brains and cultured N2a cells, while zinc chelating prevented these changes completely. Upregulation of PP2A chemically or genetically attenuated zinc-induced tau hyperphosphorylation, whereas mutation of Y307 to phenylalanine abolished the zinc-induced tyrosine phosphorylation and inactivation of PP2A. Zinc could activate Src, while PP2, a specific Src family kinases inhibitor, attenuated zinc-induced PP2A phosphorylation and inactivation, indicating that zinc induces PP2A Y307 phosphorylation and inactivation through Src activation. In human tau transgenic mice, zinc chelator rescued PP2A activity, prevented Src activation, and reduced hyperphosphorylated and insoluble tau levels. We concluded that zinc induces PP2A inactivation and tau hyperphosphorylation through Src-dependent pathway, regulation of zinc homeostasis may be a promising therapeutic for AD and the related tauopathies.     

Interleukin-7 Receptor Single Nucleotide Polymorphism rs6897932 (C/T) and the Susceptibility to Systemic Lupus Erythematosus

Emerging evidences were accumulated to support the view that aberrant interleukin-7 (IL-7) signaling might be associated with autoimmunity. Former studies demonstrated the single nucleotide polymorphism (SNP) rs6897932 C/T in the IL-7 receptor (IL-7R) gene was associated with susceptibility to autoimmune diseases, including multiple sclerosis and type I diabetes. Given these, this study was conducted to investigate whether an association existed between SNP rs6897932 and the susceptibility to systemic lupus erythematosus (SLE), a severe systemic autoimmune disease. In this context, 816 SLE patients and 816 controls from a Chinese population were recruited for this study, and the results showed that the major allele C of rs6897932 showed a higher frequency in SLE patients compared with controls (P?=?0.039, C versus T); significant difference was also detected under a recessive model with regard to the distribution of genotype frequencies between SLE patients and controls (P?=?0.041, CC versus CT + TT), which was not consistent with the results under a dominant model (P?=?0.349, CC + CT versus TT). Moreover, association studies were also performed contraposing the relationship between the SNP rs6897932 C/T and lupus nephritis as well as 10 clinical features of SLE; however, no significant association signal was found regarding the distribution of allele and genotype frequencies between SLE patients positive and negative for the presence of 11 sub-phenotypes. In conclusion, the major allele C of SNP rs6897932 may be associated with increased SLE risk in Chinese populations, and further studies are still encouraged to shed light on the true associations between SLE and its susceptibility genes with respect to IL-7R gene.     

Effect of strontium ions on the growth of ROS17/2.8 cells on porous calcium polyphosphate scaffolds

Preparation, characterization and cellular biocompatibility study of a series of calcium polyphosphate containing 0-100 mol% of Ca2+ replaced by Sr2+ were reported. The osteoblastic ROS17/2.8 cell line was used and seeded on the strontium-doped calcium polyphosphate (SCPP) scaffolds to estimate its optimal dose and to study its potential to support the growth of osteoblastic cells for bone tissue engineering. The effects of SCPP on cells' proliferation and differentiation were evaluated by MTT and ALP activity assay. The results showed that porous SCPP did not exert cytotoxic effect on the cells. In addition, the proliferation and differentiation of the growth of ROS17/2.8 cells on the SCPP containing a low dose of strontium showed a higher level compared to the control, and the SCPP containing 1% strontium was optimal according to the results of MTT and ALP activity assay. The cells on the porous SCPP formed a continuous layer on the outer and inner surface observed by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). The bunchy collagens were excreted from the cells and the calcium granules wrapped by collagens were sedimentated on the surface of cells. The results suggested that the biodegradable SCPP could stimulate the proliferation and differentiation of ROS17/2.8 cells in vitro after addition of proper dose of strontium. The porous SCPP may be a promising material for the bone tissue engineering.     

北京工科大学生需要结构调查研究

目的探讨北京工科大学的需要结构的特点。方法采用问卷法辅以访谈法,对北京某重点工科大学305名大学生进行调查。结果①工科大学生的基本需要可分为18种,其中强度最大的前4位需要依次是:友情的需要、维持生存的需要、自尊自立的需要、身体素质开发的需要;强度最弱的末4位是奉献的需要、性的需要、权力需要、减低压力的需要;②用斯皮尔曼等级相关法计算出不同年级、性别、家庭经济条件大学生等级序次之间的相关系数,结果表明不同年级(r>0.9)、性别(r=0.73)、家庭经济条件(r=0.80)的需要排列等级具有显著相关性(P<0.01);③中位数检验表明,不同年级的工科大学生在维持生存(χ2=9.12)、秩序(χ2=7.88)、自尊自立(χ2=9.81)、权力(χ2=8.36)等基本需要的强度存在显著性差异(P<0.05),在求知和求美(χ2=6.22)、精神愉悦(χ2=6.57)的基本需要的强度存在边缘显著性差异(P<0.1);不同性别的工科大学生在性(χ2=36.35)、归属(χ2=5.33)、求知求美(χ2=4.9)的基本需要强度存在显著差异(P<0.05);不同家庭经济条件的工科大学生在奉献的需要强度存在边缘显著性差异(χ2=3.49,P<0.01)。结论①北京工科大学生需要结构总体上是积极健康的,但维持生存需要强度较高,奉献需要强度需要较低;②不同年级、性别、家庭经济条件的大学生的需要结构总体上是一致的,但某些基本需要的强度存在差异。     

Hepatitis C virus-specific cellular and humoral immune responses following immunization with a multi-epitope fusion protein

Hepatitis C virus (HCV) is an important causative agent of acute and chronic hepatitis worldwide. We prepared a fusion protein in the vector of pET-11d that included three conserved broadly neutralizing B-cell epitopes and a series of T-cell epitopes located in the HCV NS3 region. In vivo administration of this fusion construct resulted in specific CD8+ cytotoxic lymphocytes in both PBMCs and splenocytes that could recognize specific antigen sites that could be detected by FACS. An HCVcc system was established and applied to detect HCV-specific neutralizing antibodies. These results suggest that the multi-epitope fusion protein is immunogenic and can elicit both humoral and cellular immune responses. In particular, this fusion protein is able to elicit HCV-specific neutralizing antibodies, which are critical for viral clearance. This construct may be significant for vaccine development and could be a potential candidate to be included in the design of a prophylactic and therapeutic vaccine against HCV.     

血管内皮生长因子C及其受体3在非小细胞肺癌组织中的表达及意义

目的　探讨血管内皮生长因子C(VEGF C)和受体 (VEGFR) 3在人非小细胞肺癌(NSCLC)组织中的表达及其与微血管、微淋巴管形成、淋巴转移、预后之间的关系。方法　对 76例人NSCLC及相应癌旁组织行VEGF C、VEGFR 3及CD34免疫组织化学染色链霉素抗生物素蛋白 过氧化物酶 (SP)法检测 ,进行淋巴管密度计数、微血管密度 (MVD)计数 ,并结合临床和病理资料进行分析。结果　NSCLC中 ,VEGF C的表达与肺癌分化程度负相关 (P =0 0 0 9)。VEGF C和VEGFR 3的表达水平与淋巴结转移呈正相关 (分别P =0 0 0 8,P =0 0 13) ,与淋巴浸润呈正相关 (分别P =0 0 2 7,P =0 0 2 0 )。VEGF C的表达与VEGFR 3在肺癌细胞中的表达呈正相关 (P =0 0 0 9)。VEGF C与淋巴管密度 (P =0 0 0 6 )、MVD(P =0 0 4 6 )呈正相关。淋巴管密度与淋巴结转移 (P =0 0 10 )、淋巴浸润 (P =0 0 19)、TNM分期 (P =0 0 15 )呈正相关 ,MVD与血行转移 (P <0 0 0 1)、TNM分期 (P <0 0 0 1)呈正相关。VEGF C阳性表达与生存时间、5年生存率呈负相关 (P <0 0 0 1)。结论　NSCLC中 ,VEGF C通过自分泌方式作用于受体VEGFR 3,促进肺癌组织生长 ,抑制分化。VEGF C促使肺癌内淋巴管形成 ,促进肺癌淋巴结转移。VEGF C和VEGFR 3表达增高、淋巴管密度增加     

Immune biology of macaque lymphocyte populations during mycobacterial infection

Immune responses of lymphocyte populations during early phases of mycobacterial infection and reinfection have not been well characterized in humans. A non-human primate model of Mycobacterium bovis bacille Calmette-Guerin (BCG) infection was employed to characterize optimally the immune responses of mycobacteria-specific T cells. Primary BCG infection induced biphasic immune responses, characterized by initial lymphocytopenia and subsequent expansion of CD4+, CD8+ and gammadelta T cell populations in the blood, lymph nodes and the pulmonary compartment. The potency of detectable T cell immune responses appears to be influenced by the timing and route of infection as well as challenge doses of BCG organisms. Systemic BCG infection introduced by intravenous challenge induced a dose-dependent expansion of circulating CD4+, CD8+ and gammadelta T cells whereas, in the pulmonary compartment, the systemic infection resulted in a predominant increase in numbers of gammadelta T cells. In contrast, pulmonary exposure to BCG through the bronchial route induced detectable expansions of CD4+, CD8+ and gammadelta T cell populations in only the lung but not in the blood. A rapid recall expansion of these T cell populations was seen in the macaques reinfected intravenously and bronchially with BCG. The expanded alphabeta and gammadelta T cell populations exhibited their antigen specificity for mycobacterial peptides and non-peptide phospholigands, respectively. Finally, the major expansion of T cells was associated with a resolution of active BCG infection and reinfection. The patterns and kinetics of CD4+, CD8+ and gammadelta T cell immune responses during BCG infection might contribute to characterizing immune protection against tuberculosis and testing new tuberculosis vaccines in primates.