SIRT2 Regulates LPS-Induced Renal Tubular CXCL2 and CCL2 Expression

Sirtuin 2 (SIRT2), a NAD⁺-dependent histone deacetylase, is involved in carcinogenesis and genomic instability and modulates proinflammatory immune responses. However, its role in renal inflammatory injury has not been demonstrated. In this study, we explored the expression patterns of CXCL2 and CCL2 in kidney tissue from Sirt2^−/− and Sirt2^+/+ mice and in mouse proximal tubular epithelial (MPT) cells. CXCL2 and CCL2 were significantly downregulated at both the mRNA and the protein levels in kidneys of LPS-treated Sirt2^−/− mice compared with those of LPS-treated Sirt2^+/+ mice. Furthermore, SIRT2 deficiency ameliorated LPS-induced infiltration of neutrophils and macrophages, acute tubular injury, and decrease of renal function. Supporting these observations, CXCL2 and CCL2 expression levels were lower in MPT cells treated with SIRT2-siRNA than in cells treated with control-siRNA, and adenovirus-mediated overexpression of SIRT2 in MPT cells significantly increased the LPS-induced expression of CXCL2 and CCL2 at the mRNA and protein levels. In addition, SIRT2 interacted with mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1), and SIRT2-knockdown increased the acetylation of MKP-1 and suppressed the phosphorylation of p38 MAPK and c-Jun N-terminal kinase in LPS-treated MPT cells. SIRT2 also regulated p65 binding to the promoters of CXCL2 and CCL2. Taken together, these findings indicate that SIRT2 is associated with expression of renal CXCL2 and CCL2 and that regulation of SIRT2 might be an important therapeutic target for renal inflammatory injury.     

重组合异种骨移植对IL－2生成影响的研究

采用重组合异种骨（ＲＢＸ）移植建立ＢＡＬＢ／Ｃ小鼠股后肌袋模型，以研究对Ｔ淋巴细胞分泌ＩＬ－２的功能变化及对细胞免疫的影响，结果显示：（１）ＲＢＸ移植后２天ＩＬ－２升高是由于手术创伤刺激所致的应激反应；（２）术后７～１４天ＲＢＸ、ｂＢＭＰ对ＩＬ－２产生有显著的抑制作用，２８～４２天回升：（３）对Ｔ淋巴细胞功能的抑制作用主要是ＢＭＰ所致，经处理的牛板质骨粒不具免疫源性。提示：ＲＢＸ移植无明显免疫排异反应，ＢＭＰ能抑制Ｔ细胞的活化增殖，使细胞因子分泌减少，因而降低厂细胞免疫水平，ＢＭＰ可能属于一种新的兔疫调节因子。实验结果对ＲＢＸ移植既能成骨又不发生明显的免疫排异的机理是一种有意义的探讨。     

二溴海因/二氧化硅复合粒子的制备及其杀菌性能评价

通过实验室制备，合成了二溴海因／二氧化硅复合粒子，运用扫描电镜（SEM）与红外光谱（IR）进行了表征，SEM结果显示，在SiO2的表面覆合了二溴海因，红外光谱图表明，复合粒子中二溴海因和二氧化硅之间以物理作用方式结合，将单纯二溴海因与二溴海因／二氧化硅复合粒子溶解于水，测定其在水中的释放速度，结果表明：二溴海因／二氧化硅复合粒子中的二溴海因在水中释放的速度明显慢于纯二溴海因，且能维持较长的作用时间，比较纯二溴海因与含等量二溴海因的复合粒子对大肠杆菌、金黄色葡萄球菌、白葡萄球菌和枯草芽孢杆菌的抑制效果，发现在二溴海因含量高于有效抑茵浓度时，复合粒子对茵体的抑制效果高于纯二溴海因。     

Multidomain synthetic peptide B2A2 synergistically enhances BMP-2 in vitro.

A multidomain, synthetic peptide designated B2A2 synergizes the activity of BMP-2. B2A2 interacts with BMP receptor isoforms, potentiating the action of BMP-2 in activating alkaline phosphatase and triggering Smad and MAPK signaling. B2A2's design permits its delivery as a local surface coating as well as a soluble co-factor, thus broadening potential bioengineering applications. INTRODUCTION: BMP-2 induces osteogenic differentiation and accelerates bone repair. Although BMP-2 inhibitors have been discovered, no BMP-2 mimetics or enhancers that function in the physiological range have yet been found. Here we report that a synthetic peptide designated B2A2, consisting of (1) a BMP receptor-targeting sequence, (2) a hydrophobic spacer, and (3) a heparin-binding sequence, is a positive modulator of recombinant BMP-2. MATERIALS AND METHODS: Cultures of mesenchymal cell lines C2C12 and C3H10T1/2 were given B2A2, recombinant BMP-2, or both. Alkaline phosphatase (ALP) activity was assayed by conversion of paranitrophenol phosphate (PNPP). Signaling through Smad and MAP kinase pathways was monitored by Western blot. Receptor binding was assessed by incubating immobilized B2A2 with soluble recombinant receptor-Fc chimeras and detecting bound receptor by anti-Fc antibody ELISA. Surface coating of medical device materials was done by first dip-coating with silyl-heparin, followed by B2A2. RESULTS AND CONCLUSIONS: Treatment of cells with B2A2 alone marginally increased ALP activity. However, B2A2 plus BMP-2 resulted in 5- to 40-fold augmentation of ALP compared with BMP-2 alone in C3H10T1/2 or C2C12 cells, respectively. This synergistic enhancement was observed over a broad concentration range (4-1000 ng/ml BMP-2). B2A2 interacted directly with BMP receptor isoforms (preferentially to BMPR-Ib and ActivinR-II). In cells, B2A2 + BMP-2 led to a repression of MAP kinase and an increase of Smad activation, consistent with known activation pathways of BMP-2. B2A2 was ineffective when paired with other cytokine/growth factors (basic fibroblast growth factor [FGF-2], TGF-beta1, vascular endothelial growth factor [VEGF]). Simultaneous co-administration was not strictly required. Pulse-chase experiments revealed that temporal separations up to 1 h were still effective. B2A2 was also effective when delivered in a polystyrene- or stainless steel-coated surface through a heparin platform (silyl-heparin) while BMP-2 was added exogenously in solution. These results suggest that B2A2 might promote aggregation of receptor subunits, enabling BMP-2 to activate signaling pathways at effectively lower concentrations. Synthetic multidomain constructs like B2A2 may be useful to accelerate bone repair/deposition through augmentation of endogenous levels of BMP-2 or through local BMP-2 contained in artificial or engineered matrices.     

PGD2-CRTH2 Pathway Promotes Tubulointerstitial Fibrosis

Urinary excretion of lipocalin-type PGD₂ synthase (L-PGDS), which converts PG H₂ to PGD₂, increases in early diabetic nephropathy. In addition, L-PGDS expression in the tubular epithelium increases in adriamycin-induced nephropathy, suggesting that locally produced L-PGDS may promote the development of CKD. In this study, we found that L-PGDS–derived PGD₂ contributes to the progression of renal fibrosis via CRTH2-mediated activation of Th2 lymphocytes. In a mouse model, the tubular epithelium synthesized L-PGDS de novo after unilateral ureteral obstruction (UUO). L-PGDS-knockout mice and CRTH2-knockout mice both exhibited less renal fibrosis, reduced infiltration of Th2 lymphocytes into the cortex, and decreased production of the Th2 cytokines IL-4 and IL-13. Furthermore, oral administration of a CRTH2 antagonist, beginning 3 days after UUO, suppressed the progression of renal fibrosis. Ablation of IL-4 and IL-13 also ameliorated renal fibrosis in the UUO kidney. Taken together, these data suggest that blocking the activation of CRTH2 by PGD₂ might be a strategy to slow the progression of renal fibrosis in CKD.Kidney failure is a public health problem worldwide, with increasing incidence and prevalence, high costs, and poor outcomes. CKD is generally progressive, incurable, and ultimately fatal, although some patients resolve with little or no sequelae. Because current treatment is basically limited to slowing the progression to ESRD using angiotensin-converting enzyme inhibitors and angiotensin II type 1 receptor blockers, more efficient therapies with different or additional modes of action are clearly needed.Regardless of disease etiology, tubulointerstitial fibrosis is the common pathway leading to ESRD in many kidney diseases and is regarded as a prognostic factor for renal function.^1–3 It is noteworthy that some clinical trials are proving that antifibrotic therapies, such as pirfenidone against diabetic nephropathy,⁴ are also effective for CKD. Therefore, elucidating the etiological mechanism underlying renal fibrosis and developing novel therapeutic strategies remains a serious, unmet medical need.Lipocalin-type PGD₂ synthase (L-PGDS) is a secretary protein of the lipocalin superfamily that converts PG H₂, a common precursor of prostanoids, to PGD₂. Because the urinary excretion of L-PGDS increases in the early stage of diabetic nephropathy,^5,6 as well as in patients with essential hypertension without any apparent renal injury,⁷ urinary L-PGDS may be an early diagnostic marker of renal injury in these patients. There is evidence indicating that, in the monkey kidney, L-PGDS is synthesized de novo in the loop of Henle, podocytes, and Bowman’s capsule of the glomeruli.⁸ Furthermore, L-PGDS gene expression in the tubular epithelium was increased in adriamycin-induced nephropathy.⁹ These findings suggest that, under conditions of tubulointerstitial stress, locally produced L-PGDS may be involved in the development of CKD. However, the precise pathophysiological significance of L-PGDS in the kidney remains to be determined.PGD₂ interacts with two receptors, the prostanoid DP₁ receptor and the chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2). Activation of the DP₁ receptor by PGD₂ has been shown to produce vasodilation¹⁰ and bronchodilation.¹¹ Furthermore, the DP₁ receptor is expressed by certain leukocyte populations,^12,13 including dendritic cells, where it controls various functions, including cytokine production. CRTH2 was originally identified as an orphan receptor expressed by Th2 lymphocytes. CRTH2 is not structurally related to the DP₁ receptor and belongs to the family of chemokine receptors. Activation of CRTH2 by PGD₂ plays an important role in allergic inflammation via the recruitment of Th2 lymphocytes and other leukocytes¹⁴ and, perhaps more importantly, by driving the production of the Th2 cytokines IL-4, IL-5, and IL-13.¹⁵