Relationship of Toll-Like Receptors 2 and 4 Gene Polymorphisms with Essential Hypertension in Chinese Han Population

Objective: There are numerous studies suggesting that genetic polymor-phisms of inflammation factors Toll-like receptors 2 and 4 (TLR2, TLR4) might play a role in the pathophysiological process of hypertension. In this study, we evaluated the association in a sample of members of the Chinese Han population. Method: We selected four single nucleotide polymor-phisms (SNP) of TLR2 (rs3804099, rs3804100, rs7656411) and TLR4 (rs1927906) genes, and measured the distributions of genotypic and allelic frequencies in 1063 participants, including 391 essential hypertension pa-tients and 672 controls. Result: No significant differences in the genotypic and allelic frequencies of the four SNPs were detected between cases and controls. However, three haplotypes, CCG, TTG and TTT of TLR2, were significantly associated with a decrease in the risk of essential hyperten-sion (OR: 0.512, 95% CI: 0.397 - 0.660, P P = 0.0038;OR: 0.797, 95% CI: 0.667 - 0.952, P = 0.0122, respectively). Inversely, the risk of essential hypertension increased sig-nificantly in patients with the CTG, TCG or TCT haplotypes (OR: 2.924, 95% CI: 2.157 - 3.963, P P P Conclusion: Our study suggested that haplotypes (CCG, TTG, TTT, CTG, TCG and TCT) of TLR2 might have profound effects on the development of essential hypertension in the Chinese Han population.

In vivo immunomodulatory profile of histamine receptors(H1,H2,H3 and H4):a comparative antagonists study

Objective:To delineate the comparative immunomodulatory roles of H1R-H4R in antibody generation profile in rabbit model.Methods:The cohort comprised of eight groups containing 18(9 male and 9 female) rabbits in each group.GroupⅠremained non-immunized and received only vehicle(sterile distilled water,1 mL/kg×b.i.d.) intramuscularly.GroupⅡreceived vehicle (1 mL/kg×b.i.d.) while GroupsⅢ-Ⅶ(drugs-treated) received subcutaneous histamine (100μg/kg×b.i.d.),and intramuscular H1R-antagonist(pheniramine,10 mg/kg×b.i.d.), H2R-antagonist(ranitidine,10 mg/kg×b.i.d.),H3R-antagonist(iodophenpropit,1μg/kg×b.i.d.) and H4R-antagonist(JNJ 7777120,10μg/kg×b.i.d.),and GroupⅧDMSO(1 mL/kg×b.i.d.),respectively for 10 days(starting from day 1).They were subsequendy immunized with intravenous injection of sheep red blood cells(SRBC) at day 3.The estimation of serum Igs,IgM and IgG were done by ELISA,and observed at day 0(pre-immunization),and 7,14,21,28 and 58(post-immunization).Results:It was shown that histamine and HRs-antagonists could influence a detectable antibody response to SRBC as early as day 7-post-immunization(post-Ⅰ), which lasted until day 58 post-Ⅰ.The results were found statistically significant(P【0.05,). Conclusions:This study suggests that histamine receptors play important roles in modulation of antibody generation in which H1R,H2R and H4R have immunosuppressive roles and conversely, H3R playes an immune enhancing role.The findings of this study may have clinical significance and provide the baseline information for future study.

Anti-Anxiety Effects of Prelimbic 5-HT4 Receptors in the Rat Model of Parkinson’s Disease: Evidence from Behavioral and Electrophysiological Study

Objective:Clinical and laboratory studies have demonstrated that prelimbic(PrL)and serotonin-4(5-HT4)receptors may have the key role in regulating anxiety.However,the pathophysiology of anxiety in Parkinson’s disease(PD)remains obscure.In this research,the effects of PrL 5-HT4 receptors on anti-anxiety behaviors in hemiparkinsonian rats were investigated.Methods:PD model rats were used as the research subjects,starting with behavioral changes,from the point of view of electrophysiology,the regulatory effect of PrL 5-HT4 receptors on PD-related anxiety and the possible mechanism were explored.Results:Anxiety-like behaviors were induced via MFB lesion in rats.Intra-PrL injection of 5-HT4 receptors agonist RS67333 induced anti-anxiety effects in both sham and PD group.In the sham group,PrL administration of 5-HT4 receptors antagonist SB204070 produce anti-anxiety effects,but in the PD group,the expression of anxiety-like behavior was increased.Compared to the sham group,the effective dose of the behavioral effects of the two drugs in the PD group was obviously higher.Electrophysiological data suggested that PrL administration of RS67333(SB204070)increased(decreased)the firing activities ofγ-aminobutyric acid(GABA)neurons in both groups.Compared with rats in sham group,lesioned rats had a shorter duration of the excitation(inhibition)effects on firing activities of GABA neurons.Conclusion:PrL 5-HT4 receptors regulate anxiety behaviors in PD rats,and its mechanism may be related to the down-regulation of expression or function of PrL 5-HT4 receptors in PD.

Divergent expression of bacterial wall sensing toll-like receptors 2 and 4 in colorectal cancer

AIM To characterize the expression of toll-like receptors(TLR) 2 and 4 in colorectal cancer(CRC) and in normal colorectal mucosa.METHODS We analysed tissue samples from a prospective series of 118 unselected surgically treated patients with CRC. Sections from formalin fixed, paraffin embedded specimens were analysed for TLR2 and TLR4 expression by immunohistochemistry. Two independent assessors evaluated separately expression at the normal mucosa, at the invasive front and the bulk of the carcinoma, and in the lymph node metastases when present. Expression levels in different locations were compared and their associations with clinicopathological features including TNM-stage and the grade of the tumour and 5-year follow-up observations were analysed.RESULTS Normal colorectal epithelium showed a gradient of expression of both TLR2 and TLR4 with low levels in the crypt bases and high levels in the surface. In CRC, expression of both TLRs was present in all cases and in the major proportion of tumour cells. Compared to normal epithelium, TLR4 expression was significantly weaker but TLR2 expression stronger in carcinoma cells. Weak TLR4 expression in the invasive front was associated with distant metastases and worse cancerspecific survival at 5 years. In tumours of the proximal colon the cancer-specific survival at 5 years was 36.9% better with strong TLR4 expression as compared with those with weak expression(P = 0.044). In contrast, TLR2 expression levels were not associated with prognosis. Tumour cells in the lymph node metastases showed higher TLR4 expression and lower TLR2 expression than cells in primary tumours.CONCLUSION Tumour cells in CRC show downregulation of TLR4 and upregulation of TLR2. Low expression of TLR4 in the invasive front predicts poor prognosis and metastatic disease.

TypeⅠinositol 1, 4, 5-triphosphate receptors increase in kidney of mice with fulminant hepatic failure

AIM: To delineate the mechanisms of renal vasocon- striction in hepatorenal syndrome (HRS), we investigated the expression of typeⅠinositol 1, 4, 5-triphosphate receptors (IP3RⅠ) of kidney in mice with fulminant hepatic failure (FHF). METHODS: FHF was induced by lipopolysaccharide (LPS) in D-galactosamine (GalN) sensitized BALB/c mice. There were 20 mice in normal saline (NS)-treated group, 20 mice in LPS-treated group, 20 mice in GalN- treated group, and 60 mice in GalN/LPS-treated group (FHF group). Liver and kidney tissues were obtained at 2, 6, and 9 h after administration. The liver and kidney specimens were stained with hematoxylin-eosin for studying morphological changes under light microscope. The expression of IP3RⅠin kidney tissue was tested by immunohistochemistry, Western blot and reverse transcription (RT)-PCR. RESULTS: Kidney tissues were morphologically normal at all time points in all groups. IP3RⅠproteins were found localized in the plasma region of glomerular mesangial cells (GMC) and vascular smooth muscle cells (VSMC) in kidney by immunohistochemical staining. In kidney of mice with FHF at 6 h and 9 h IP3RⅠstaining was up- regulated. Results from Western blot demonstrated consistent and significant increment of IP3RⅠexpression in mice with FHF at 6 h and 9 h (t = 3.16, P < 0.05; t = 5.43, P < 0.01). Furthermore, we evaluated IP3RⅠ mRNA expression by RT-PCR and observed marked up- regulation of IP3RⅠmRNA in FHF samples at 2 h, 6 h and 9 h compared to controls (t = 2.97, P < 0.05; t = 4.42, P < 0.01; t = 3.81, P < 0.01). CONCLUSION: The expression of IP3RⅠprotein increased in GMC and renal VSMC of mice with FHF, possibly caused by up-regulation of IP3RⅠmRNA.

M_(4) muscarinic receptors regulates dopamine/DARPP-32 signaling and glutamate transmis⁃sion to balance dopaminergic D1 function in mouse dorsal striatum

OBJECTIVE Abnormal striatal dopaminergic and glutamatergic neurotransmis⁃sion is central to the pathophysiology of schizo⁃phrenia.In this study,we investigated the roles of M4 receptor interplay with D1 signaling in stria⁃tal neurotransmission that affect glutamatergic transmission to control the etiology of neuropsy⁃chiatric disorders.METHODS To study dorsal striatum(DS)region-specific neuronal and behav⁃ioral responses modulated by M4 receptors,we used clustered regularly interspaced short palin⁃dromic repeats-associated protein 9 technology to generate mice lacking M4 in the dorsal stria⁃tum(DS-M4-KD).The M4 positive allosteric modu⁃lator,VU0467154,were used to study the phar⁃macologically profiles with M4 receptor stimula⁃tion in WT mice.Oxotremorine M(Oxo-M),a no subtype-selective muscarinic agonist,was used to show that mAchRs activation,in order to dissect the particular function of M4,in DS-M4-KD mice.Open filed test and forced swim test were used to assess the change of psychiatric-like behav⁃iors.Western blotting and immunohistochemistry were used to detect protein levels of phosphory⁃lation site of dopamine-and cAMP-regulated phosphoprotein of 32 ku(DARPP-32).Whole-cell patch-clamp recording was used to assess M4-mediated cholinergic inhibition of glutamater⁃gic synaptic input transmission.RESULTS West⁃ern blotting and immunohistochemistry assay showed VU0467154(5 mg·kg-1,ip)promoted phosphorylation of DARPP-32 at Thr75,and atten⁃uated D1-dependent phosphorylation of DARPP-32 at Thr34 within the mouse DS.Consistently,the Oxo-M(4μg,icv)also increased DARPP-32 phosphorylation at site Thr75 to reversed phos⁃phorylation at site Thr34 in WT mice,but not in DS-M4-KD mice.In parallel with altered DARPP-32 responses,VU0467154 or Oxo-M evoked a psychological stress response and reversed D1-induced hyperlocomotion in mice in open field test and force swim tests.However,Oxo-M sup⁃pression of D1-depengdeng behavioral respons⁃es was impaired in DS-M4-KD mice.Whole-cell patch recording showed that VU0467154 or Oxo-M mediated endogenous cholinergic inhibition of miniature excitatory postsynaptic currents through M4 receptors,which in turn suppressed D1-depen⁃dent glutamatergic synaptic transmission in the DS.CONCLUSION This study provides evidence for the role of M4 receptors in regulation of dopa⁃mine/DARPP-32 signaling and glutamate respons⁃es in the DS,and therefore modulation of psychi⁃atric behaviors associated with D1 signaling.This results indicate the mechanisms of treatments targeting M4 in psychiatric disorders.

CXCL12 Retargeting of an Oncolytic Adenovirus Vector to the Chemokine CXCR4 and CXCR7 Receptors in Breast Cancer

Breast cancer is the most frequently diagnosed cancer in women under 60, and the second most diagnosed cancer in women over 60. While significant progress has been made in developing targeted therapies for breast cancer, advanced breast cancer continues to have high mortality, with poor 5-year survival rates. Thus, current therapies are insufficient in treating advanced stages of breast cancer;new treatments are sorely needed to address the complexity of advanced-stage breast cancer. Oncolytic virotherapy has been explored as a therapeutic approach capable of systemic administration, targeting cancer cells, and sparing normal tissue. In particular, oncolytic adenoviruses have been exploited as viral vectors due to their ease of manipulation, production, and demonstrated clinical safety profile. In this study, we engineered an oncolytic adenovirus to target the chemokine receptors CXCR4 and CXCR7. The overexpression of CXCR4 and CXCR7 is implicated in the initiation, survival, progress, and metastasis of breast cancer. Both receptors bind to the ligand, CXCL12 (SDF-1), which has been identified to play a crucial role in the metastasis of breast cancer cells. This study incorporated a T4 fibritin protein fused to CXCL12 into the tail domain of an adenovirus fiber to retarget the vector to the CXCR4 and CXCR7 chemokine receptors. We showed that the modified virus targets and infects CXCR4- and CXCR7-overexpressing breast cancer cells more efficiently than a wild-type control vector. In addition, the substitution of the wild-type fiber and knob with the modified chimeric fiber did not interfere with oncolytic capability. Overall, the results of this study demonstrate the feasibility of retargeting adenovirus vectors to chemokine receptor-positive tumors.

Role of liver X receptors alpha agonist on expressions of LPS-induced inflammatory response associated factor IRAK-4 and NF-kappaB in Kupffer cells

在 interleukin-1 受体的表情上探索激活的肝 X 受体(LXR ) 的角色的目的在在 Kupffer 房间并且到由 LPS 导致了的煽动性的反应联系了 kinase-4 (IRAK-4 ) 和 NF-kappaB (NF-B ) 调查煽动性的反应的 LXR 否定规定的可能的机制。Kupffer 房间被骨胶原灌注在 situ 从男 Kunming 老鼠孤立的方法。并且这些房间被划分成 4 个组：正常控制组， LPS 处理组， LXR 收缩筋 T0901317 处理组， LPS 和 T0901317 联合了处理组。 LPS 处理组在 RPMI 1640 与 1 g/ml LPS 的最后的集中被对待并且为 6 h 有教养， T0901317 处理组在 RPMI 1640 与 5 g/ml 的最后的集中被对待并且为 24 h 有教养，并且联合处理组在 RPMI 1640 与 1 g/ml T0901317 的最后的集中为 24 h 收到了文化前然后为有在 RPMI 1640 的 5 g/ml LPS 的最后的集中的 6 h 有教养。所有组为 30 h 是有教养的。在 mRNA 和蛋白质层次的 LXR， IRAK-4 和 NF-B 的表示被即时 PCR 并且西方的弄污检测，并且 TNF-1 和 IL-1 层次被 ELISA 检测。LXR mRNA 和蛋白质的层次是的结果在 LPS 组在 T0901317 组最高、最低(P < 0.05 ) 。IRAK4 和 NF-B mRNAs 和蛋白质的水平比在 LPS 组在联合对待的组是显然更低的(P < 0.05 ) 。并且 TNF-1 和 IL-1 的水平在 LPS 组最高被观察(P < 0.05 ) ，但是在控制组， T0901317 组和联合对待的组之中的没有差别(P > 0.05 ) 。这些标明日期的结论建议 LXR 收缩筋有效地装起来调整 LXR mRNA 和蛋白质的表情并且禁止煽动性的反应。这可能经由下面调整在 mRNA 和蛋白质层次的 IRAK4 和 NF-B 的表情。

Expression of BMP Receptors in Porcine Granulosa Cells (GCs) and Their Regulation by Luteinizing Hormone (LH)

Bone morphogenetic proteins（BMPs） play critical roles in follicle growth and development.BMPs initiate signaling by assembling BMP receptors and activating Smads,which in turn alter expression of target genes.The mechanism underlying the regulation of the expression of BMP receptors and Smads during follicle development in pigs is still unknown.By quantitative real-time PCR,the mRNA expression of BMP receptors and Smads in granulosa cells（GC） was investigated.Cells were obtained from small porcine follicles（SF;3 mm diameter） and dominant follicles（DF;6 mm diameter）;ActR1A and BMPR2 mRNA levels in DF were significantly higher（P0.05） than that in SF,whereas BMPR1B,Smad4 and Smad7 expression tended to decrease（P0.05）.The levels of BMPR1A,ActR2,Smad1,Smad5,and Smad8 mRNA did not differ between DFs and SFs.To investigate the effect of LH on BMP receptors in GC,cells obtained from porcine DFs were cultured in medium supplemented with different doses of luteinizing hormone（LH）.High doses of LH（4 IU mL-1） significantly decreased the concentration of estradiol（E2） and progesterone（P4） in medium and the expression of Cyp19a1（P450 aromatase,P450arom） and Cyp11a1（cholesterol side-chain cleavage enzyme P450,P450scc）,while significantly increased viable cell numbers and up-regulated expression of cyclin dependent kinase-4（CDK4） and cyclin D2.However,LH had no effect on the expression of BMP receptor genes.Thus,the present study indicates that the expression of members of the BMP signaling pathway in porcine GC is regulated during follicle development and the expression of BMP receptors are not regulated by LH in porcine GCs.

Uridine adenosine tetraphosphate acts as a pro-angiogenic factor in vitrothrough purinergic P2Y receptors

OBJECTIVE Uridine adenosine tetraphosphate(Up4A),a dinucleotide,contains both purine and pyrimidine moieties,and exerts its vascular influence via activation of purinergic receptors.Here,we aimed to investigate the effects of Up4 A on angiogenesis and the putative purinergic receptors(PR)involved in this process.METHODS Tubule formation assay was performed in 3D matrix system.In this assay,human umbilical vein endothelial cells(HUVECs)were co-cultured with pericytes with various Up4 A doses(0,1,2.5,5,10 and 20μmol·L-1)in the absence and presence of P2Y6 R antagonist MRS2578(10μmol·L-1)for 5d.Expression profile of PR subtypes and angiogenic factors was assessed in HUVECs by q-PCR with and without P2Y6 R antagonist.RESULTS No difference in initial tubule formation was detected between Up4 A stimulation and control conditions at day 2.In contrast,a significant increase in vascular density in response to Up4 A was observed at day 5.Up4 A at a dose of 2.5and 5μmol·L-1 promoted total tubule length(by-1.89 fold and-2.23fold),number of tubules(by-1.71 fold and-1.89fold)as well as number of junctions(by-2.24 fold and-2.80fold),all of which were inhibited by MRS2578.Further increase in Up4 A dose to10 and 20μmol·L-1 did not induce an increase in these vascular parameters as compared to non-treated controls.Moreover,Up4 A increased mRNA level of P2YRs(P2Y2R,P2Y4 R and P2Y6R)but not P2XR(P2X4R and P2X7R)or P1R(A2AR and A2BR),while Up4 A upregulated VEGFA and ANGPT1 but not VEGFR2,ANGPT2,Tie1 and Tie2at mRNA level.Transcriptional upregulation of P2 YRs and angiogenic factors by Up4 A was inhibited by MRS2578.CONCLUSION Up4 A is functionally capable of promoting tubule formation in vitro co-culture system.This process is likely mediated by activation of pyrimidine-favored P2 YRs but not P2 XR or P1 Rs,and involves stimulation of well known angiogenic factors.

3-Arylisothiazoloquinols as potent ligands for the benzodiazepine site of GABA_(A) receptors

3-Arylisothiazolo[5,4-b]quinolin-4(9H)-ones and 3-arylisoxazolo[5,4-b]quinolin-4(9H)-ones were synthesized and assayed for affinity for the benzodiazepine binding site of the GABAA receptors. While the 3-arylisothiazoloquinolin-4-ones were found to be potent ligands, with affinities (expressed as the affinity Ki value) down to 1 nM, the 3-arylisoxazoloquinolin-4-ones are less potent. This is suggested to depend on sterical repulsive interaction of the 3-arylisoxazoloquinolin-4-ones with the receptor essential volume of the binding site, and a higher electron density at the nitrogen in the azole ring (N-2) as well as the carbonyl oxygen in the isothiazoloquinolin-4-ones enabling them to interact stronger with hydrogen bond donor sites at the binding site.

Association between Promoter Polymorphisms of Interleukin-4 Gene and Allergic Rhinitis Risk: a Meta-analysis

Summary： The relationship of interleukin-4 （IL-4） C-33T and C-590T （C-589T） gene polymorphisms with allergic rhinitis was analyzed. Data about the case control studies of IL-4 gene promoter polymorphisms [C-33T and C-590T （C-589T）] and their association with allergic diseases and correlation between serum IL-4 levels and allergic rhinitis were retrieved. The Stata 12.0 statistical soitvcare was applied to analyze the correlation between IL-4 gene polymorphisms and allergic rhinitis. The meta-analysis result of TT/CC genotype of -590 （-589） polymorphism showed a significant association with allergic diseases [OR=1.93, 95% CI （1.61 2.31）, P=0.00]. Meta-analysis of the TT＋TC versus CC genotype of IL-4 C-33/T polymorphism revealed significant associations with allergic diseases [OR=3.23, 95% CI （1.13-9.25）, P=0.03]. Meanwhile, there was a significant correlation between serum IL-4 levels and allergic rhinitis [OR=2.52, 95% CI-（1.80-3.23）, P=0.00]. IL-4 gene -590 TT genotype may increase the risk of allergic rhinitis and the T allele mutation of -33 might be correlated with aller- gic rhinitis.

Role of P2X_7 receptors in the development of diabetic retinopathy

The P2X7 receptor is one of the members of the family of purinoceptors which are ligand-gated membrane ion channels activated by extracellular adenosine 5'-triphosphate. A unique feature of the P2X7 receptor is that its activation can result in the formation of large plasma membrane pores that allow not only the flux of ions but also of hydrophilic molecules of up to 900 Da. Recent studies indicate that P2X7-mediated signaling can trigger apoptotic cell death after ischemia and during the course of certain neurodegenerative disorders. Expression of the P2X7 receptor has been demonstrated in most types of cells in the retina. This purinoceptor mediates the contraction of pericytes and regulates the spatial and temporal dynamics of the vasomotor response through cell-to-cell electrotonic transmission within the microvascular networks. Of potential clinical significance, investigators have found that diabetes markedly boosts the vulnerability of retinal microvessels to the lethal effect of P2X7 receptor activation. This purinergic vasotoxicity may result in reduced retinal blood flow and disrupted vascular function in the diabetic retina. With recent reports indicating an association between P2X7 receptor activation and inflammatory cytokine expression in the retina, this receptor may also exacerbate the development of diabetic retinopathy by a mechanism involving inflammation.

The Relationship between Polymorphisms of Interleukin-4 Gene and Silicosis

Objective To explore the relationship between polymorphisms of interleukin-4 （IL-4） gene （-33, ＋45, intron3, ＋429, ＋448） and the susceptibility of silicosis. Methods A case-control study was carried out. 101 silicosis patients were selected as cases. As strictly matching, 121 of non silicosis workers were selected as the controls. The polymophisms of IL-4 （five locus） were detected by the method of polymerase chain reaction-restriction fragment length polymorphism （PCR-RFLP） techniques. Results The GA genotype in the IL-4＋429 locus and the CC genotype in the IL-4＋448 locus were found. The frequencies ofAA, GG and AG of IL-4＋45 locus in the cases were 55.4%, 10.9%, and 33.7% and in the controls were 62.0%, 12.6%, and 26.4%. The differences between cases and controls were not significant. The frequencies of B1B1, B2B2, and B1B2 of intron3 VNTR locus in the cases were 73.3%, 1.0%, and 25.7% and in the controls were 68.6%, 1.7%, and 29.8%. The differences were not significant. The frequencies of TT, CC, and CT in -33 locus in the cases were 55.4%, 11.9%, and 32.7% and in the controls were 69.4%, 4.1%, and 26.4%. The differences were significant （P=0.034）. Conclusion The relationship between genetic polymorphism of IL-4-33 site and silicosis has been found and -33TT is a protective genotype for silicosis.

Effect of ω-3 Polyunsaturated Fatty Acid on Toll-like Receptors in Patients with Severe Multiple Trauma

This study examined the effects of ω-3 polyunsaturated fatty acid（ω-3PUFA） on the expression of toll-like receptor 2（TLR2）,toll-like receptor 4（TLR4） and some related inflammatory factors in peripheral blood mononuclear cells（PBMCs） of patients with early-stage severe multiple trauma.Thirty-two patients who were admitted to the Department of Traumatic Surgery,Tongji Hospital（Wuhan,China） between May 2010 and November 2010,and diagnosed as having severe multiple trauma with a injury severity score（ISS） no less than 16,were enrolled in the study and divided into two groups at random（n=16 in each）：ω-3PUFA group and control group in which routine parenteral nutrition supplemented with ω-3PUFA or not was administered to the patients in two groups for consecutive 7 days.Peripheral blood from these patients was collected within 2 h of admission（day 0）,and 1,3,5 and 7 days after the nutritional support.PBMCs were isolated and used for detection of the mRNA and protein expression of TLR2 and TLR4 by using real-time PCR and flow cytometry respectively,the levels of NF-κB by quantum dots-based immunofluorescence assay,the levels of TNF-α,IL-2,IL-6 and COX-2 by ELISA,respectively.The results showed that the mRNA and protein expression of TLR2 and TLR4 in PBMCs was significantly lower in ω-3PUFA group than in control group 5 and 7 days after nutrition support（both P0.05）.The levels of TNF-α,IL-2,IL-6 and COX-2 were found to be substantially decreased in PBMCs in ω-3PUFA group as compared with control group at 5th and 7th day（P0.05 for all）.It was concluded that ω-3PUFA can remarkably decrease the expression of TLR2,TLR4 and some related inflammatory factors in NF-κB signaling pathway in PBMCs of patients with severe multiple trauma,which suggests that ω-3PUFA may suppress the excessive inflammatory response meditated by the TLRs/NF-κB signaling pathway.

The Akt/glycogen synthase kinase-3β pathway participates in the neuroprotective effect of interleukin-4 against cerebral ischemia/reperfusion injury

Interleukin-4(IL-4) has a protective effect against cerebral ischemia/reperfusion injury. Animal experiments have shown that IL-4 improves the short-and long-term prognosis of neurological function. The Akt(also called protein kinase B, PKB)/glycogen synthase kinase-3β(Akt/GSK-3β) signaling pathway is involved in oxidative stress, the inflammatory response, apoptosis, and autophagy. However, it is not yet clear whether the Akt/GSK-3β pathway participates in the neuroprotective effect of IL-4 against cerebral ischemia/reperfusion injury. In the present study, we established a cerebral ischemia/reperfusion mouse model by middle cerebral artery occlusion for 60 minutes followed by a 24-hour reperfusion. An IL-4/anti-IL-4 complex(10 μg) was intraperitoneally administered 30 minutes before surgery. We found that administration of IL-4 significantly alleviated the neurological deficits, oxidative stress, cell apoptosis, and autophagy and reduced infarct volume of the mice with cerebral ischemia/reperfusion injury 24 hours after reperfusion. Simultaneously, IL-4 activated Akt/GSK-3β signaling pathway. However, an Akt inhibitor LY294002, which was injected at 15 nmol/kg via the tail vein, attenuated the protective effects of IL-4. These findings indicate that IL-4 has a protective effect on cerebral ischemia/reperfusion injury by mitigating oxidative stress, reducing apoptosis, and inhibiting excessive autophagy, and that this mechanism may be related to activation of the Akt/GSK-3β pathway. This animal study was approved by the Animal Ethics Committee of Renmin Hospital of Wuhan University, China(approval No. WDRY2017-K037) on March 9, 2017.

Role of toll receptors in diabetic nephropathy

Objectives: Diabetic nephropathy is the leading cause of chronic kidney disease. The pathogenesis of DN remains incompletely understood. It has been recently demonstrated that inflammatory processes play a significant role in the development and progression of DN. Toll-like receptors play a fundamental role in the innate immune system by triggering proinflammatory signaling pathways. Our aim is to evaluate the expression of TLRs on monocytes and relate their expression with inflammation in HD patients with & without diabetic nephropathy. Method: In a case control study (60) patients from Alkasr El Aini Hospital on hemodialysis were divided into two groups: Group 1, 30 patients on heamodialysis not due to diabetic nephropathy, Group 2, 30 patients on heamodialysis due to diabetic nephropathy, compared to Group 3, including 30 healthy controls. All participants were subjected to: Full medical history, complete physical examination, Serum creatinine, uric acid, A1C, fundus examination, detection of TLR2, TLR expression by real time PCR in peripheral blood mononuclear cells. Data were statically calculated using SPSS, comparision between groups was done using student T test comparing 2 groups, correlation using spearman’s correlation. Results: Diabetic had significantly increased TLR2, TLR4 mRNA in peripheral blood mononuclear cells compared to controls and non diabetics patient on heamodialysis (p < 0.001), TLR2, TLR4 significantly correlated with dialysis duration in diabetic (p < 0.001), no correlation with A1C in relation to TLR2 (p = 0.078), TLR4 (p = 0.163). Conclusion: TLR2, TLR4 were significantly elevated in diabetic on dialysis initiating event in the pathogenesis of DN, providing a link between hyperglycemia and hypoxia with inflammation and fibrosis within the kidney. Hence, therapeutic interventions aimed at targeting the inflammatory component through interruption of TLR signaling may be a novel strategy to target prevention and treatment of DN.

Effect of neuropeptide Y on white matter demyelination and serum interleukin-4 and gamma-interferon levels in the guinea pig with experimental allergic encephalomyelitis

BACKGROUND： Neuropeptide Y （NPY） may influence differentiation of Th cells immunological pathology of experimental allergic encephalomyelitis （EAE） is differentiation of Th cells It is assumed that the related to abnormal OBJECTIVE： To investigate the effect of NPY on white matter demyelination, the serum levels interleukin-4 （IL-4） and gamma-interferon （IFN-γ ）, as well as EAE pathogenesis in an EAE guinea pig model following NPY injection into the lateral cerebral ventricle. DESIGN, TIME AND SETTING： A randomized controlled animal study, which was performed in the Infection Immunity Animal Laboratory, Affiliated Hospital of Luzhou Medical College, China, from October 2005 to April 2006. MATERIALS： Thirty healthy female guinea pigs of 8-12 weeks of age, and 10 healthy female rats of three months of age were used. NPY was provided by Sigma Company, USA. NPY kit was provided by Beijing Huaying Biotechnology Institute, China. METHODS： Thirty guinea pigs were randomly divided into three groups： normal control group, EAE model group, and NPY intervention group （n =10 per group）. Normal control group and EAE model group： Saline （10μ L, once） was injected into the lateral cerebral ventricle. After one week, the same volume of Freund＇s adjuvant complete was either injected subcutaneously into two post-palms or EAE was modeled. NPY intervention group： EAE was modeled after one week and NPY was injected （10 μ L of 6 nmol NPY, once） into the lateral cerebral ventricle. Myelin basic protein （MBP） antigen made from rat spinal cord homogenate and Freund＇s adjuvant complete were injected subcutaneously into both post-palms （0.2 mL per palm） to establish the EAE model. MAIN OUTCOME MEASURES： White matter demyelination of the cerebrum, cerebellum, brain stem, and spinal cord were observed by light microscopy after HE staining. Levels of serum IFN-γ and IL-4 were detected by the double antibody sandwich ABC-ELISA technique. NPY content was detected by radioimmunoassay. RESULTS： Pathological alterations in the NPY intervention groups were reduced compared to those in the EAE model group, suggesting a reduction and remission of white matter demyelination with NPY treatment. When compared to the model group, the serum IL-4 level was increased in the NPY intervention group during the high-frequent EAE stage （P 〈 0.01）, but the serum IFN-γ level was decreased （P 〈 0.01）. Furthermore, the EAE latency was prolonged （P 〈 0.01）, the neurological scores were decreased in the high-frequent EAE stage （P 〈 0.01）, and the death rate was decreased （P 〈 0.05）. NPY content and the serum IL-4 level at the peak stage were positively correlated with those in the latent phase （r =0.863-0.900, P 〈 0.01）, but negatively correlated with neurological scores at the peak stage （r=- -0.068 to -0.863, P 〈 0.05-0.01）. The IFN-γ level at the peak stage was negatively correlated to that in the latent phase （r = -0.683-0.650, P 〈 0.05）, but positively correlated to neurological scores at the peak stage （r =0.975, 0.845, P 〈 0.05）. CONCLUSION： NPY injection into the lateral cerebral ventricle can promote the secretion of IL-4, inhibit the production of IFN-γ, relieve white matter demyelination, and inhibit EAE attack in an experimental model of EAE.

Effects of Intrinsic Nitric Oxide on the Expression of Interleukin-4 and IFN-γ mRNA in the Bronchial and Lung Tissues of Sensitized Rats

Summary: To investigate the effects of intrinsic nitric oxide (NO) on the expression of interleukin-4 (IL-4) mRNA and interferon-γ (IFN-γ) mRNA in the airway inflammation of asthma, the rat models of asthmatic inflammation were established by sensitizing and then challenging the animals with ovalbumin. The 24 animals were randomly divided into control group, sensitized group, sensitized and L-Arg-treated group as well as L-NAME-treated group equally. By using in situ hybridization combined with compute physiological quantitative imaging analysis techniques, the influence of intrinsic NO on the expression of IL-4 mRNA and IFN-Y mRNA in the airway inflammatory cells was observed. In situ hybridization study demonstrated that IL-4 mRNA expres- sion was obviously increased as compared with that in the control group, mainly distributed in the inflammatory cells in the submucous of airways in the sensitized group. The increase of intensity of IL-4 mRNA expression was positively correlated with the numbers of eosinophil (Eos) and lymphocyte (both with P<0. 05) in the sensitized group. There was no statistically difference IFN- γ expression between the control group and the sensitized group. Imaging analysis showed that L- NAME could inhibit the expression of IL-4 mRNA (P<0. 05) and increase the expression of IFNY mRNA (P<0. 05), while L-Arg could increase the expression of IL-4 mRNA in inflammatory cells (P<0. 05). It was indicated that a suitable levels of intrinsic NO can influence the expression of IL-4 mRNA of Th2 lymphocytes and the expression of IFN-γ mRNA of Th1 lymphocytes and in turn, promote the development of asthmatic airway inflammation.

Interactions of thymic stromal lymphopoietin with interleukin-4 in adaptive immunity during Aspergillus fumigatus keratitis

AIM:To investigate the potential interactions of thymic stromal lymphopoietin(TSLP)with interleukin-4(IL-4)in adaptive immunity during fungal keratitis(FK).METHODS:An FK mouse model was induced with Aspergillus fumigatus(AF)hyphal infection.Mice were divided into several groups:untreated,phosphate buffer saline(PBS),infected with AF,and pretreated with a scrambled siRNA,a TSLP-specific siRNA(TSLP siRNA),murine recombinant TSLP(rTSLP),immunoglobulin G(IgG),murine recombinant IFN(rIFN-γ),murine recombinant IL-4(rI L-4),rIL-13,murine recombinant IL-17A(rIL-17A),and murine recombinant IL-17F(rIL-17F)groups.Quantitative realtime reverse transcription-polymerase chain reaction(qRTPCR)and enzyme-linked immunosorbent assay(ELISA)or Western blot were performed to determine mRNA and protein levels in the inflamed cornea.Cytokine locations were observed by immunofluoresence staining after AF hyphal infection.RESULTS:Compared to those in the untreated group,TSLP and T helper type 1(Th1)cytokine levels in the AF group were upregulated at 24 h post infection(hpi),and those of T helper type 2(Th2)and T helper type 17(Th17)cytokines were increased at 5 d post infection(dpi).Th2 cytokine levels were decreased in the TSLP siRNA-pretreated group and increased in the rTSLP-pretreated group compared with the AF group.The TSLP level was increased in the rIL-4-pretreated group,but there were no significant changes among the other groups.Immunofluorescence staining showed cytokine locations after AF hyphal infection.CONCLUSION:TSLP induces a Th2 immune response and promots Th2 T cell differentiation in vivo.IL-4 promotes TSLP secretion.Therefore,TSLP with IL-4 regulates adaptive immunity in FK.