Monocyte Chemotactic Protein-1 mRNA Research Articles

Sodium chloride promotes tissue inflammation via osmotic stimuli in subtotal-nephrectomized mice

Chronic inflammation, which is often associated with high all-cause and cardiovascular mortality, is prevalent in patients with renal failure; however, the precise mechanisms remain unclear. High-salt intake was reported to induce lymphangiogenesis and autoimmune diseases via osmotic stimuli with accumulation of sodium or chloride. In addition, sodium was recently reported to be stored in the extremities of dialysis patients. We studied the effects and mechanisms of high salt loading on tissue and systemic inflammation in subtotal-nephrectomized mice (5/6Nx) and in cultured cells. Macrophage infiltration in the peritoneal wall (P<0.001), heart (P<0.05) and para-aortic tissues (P<0.001) was significantly higher in 5/6Nx with salt loading (5/6Nx/NaCl) than in 5/6Nx without salt loading (5/6Nx/Water); however, there were no significant differences in blood pressure and renal function between the groups. Tissue interleukin-6, monocyte chemotactic protein-1 (MCP-1), serum- and glucocorticoid-inducible kinase 1 (Sgk1) and tonicity-responsive enhancer binding protein (TonEBP) mRNA were significantly elevated in the peritoneal wall and heart with 5/6Nx/NaCl when compared with 5/6Nx/Water. Sodium was stored in the abdominal wall, exerting high-osmotic conditions. Reversal of salt loading reduced macrophage infiltration associated with decreased TonEBP in 5/6Nx/NaCl. Macrophage infiltration associated with fibrosis induced by salt loading was decreased in the 5/6Nx/NaCl/CC chemokine receptor 2 (CCR2, receptor of MCP-1)-deficient mice when compared with 5/6Nx/NaCl/Wild mice, suggesting that CCR2 is required for macrophage infiltration in 5/6Nx with NaCl loading. In cultured mesothelial cells and cardiomyocytes, culture media with high NaCl concentration induced MCP-1, Sgk1 and TonEBP mRNA, all of which were suppressed by TonEBP siRNA, indicating that both MCP-1 and Sgk1 are downstream of TonEBP. Our study indicates that high NaCl intake induces MCP-1 expression leading to macrophage infiltration via the TonEBP-MCP-1 pathway in 5/6Nx/NaCl mice, and that TonEBP has a central role in inflammation in patients with renal failure taking high salt.

Abstract A29: Effects of obesity on regulation of macrophage population in the prostate tumor microenvironment

Abstract Background: Obesity is associated with a greater risk of prostate cancer mortality, but the mechanisms driving this progression are still unclear. A high concentration of macrophages within the tumor microenvironment, especially tumor-associated macrophages (TAMs), has been linked to progression and metastasis in numerous cancer types, including prostate. Obesity increases circulating levels of monocyte chemotactic protein-1 (MCP-1), a chemokine involved in monocyte migration. In addition, prostaglandin E2 (PGE2), a molecule closely associated with obesity, appears to be involved in macrophage polarization. Our preliminary in vitro data suggest that obese conditions promote both macrophage recruitment and TAM polarization. Hypothesis: Based upon these preliminary results, we hypothesize that obesity-induced prostate cancer progression is due, in part, to increased TAM concentrations in the prostate tumor micro-environment, promoting pro-metastatic signaling. Methods: In order to mimic the prostate tumor microenvironment, an in vitro model composed of adipose stromal cells, macrophages, prostate cancer epithelial cells and sera from obese and non-obese men was used to assess the impact of obesity on the paracrine signaling network. Matrigel invasion chambers were used to assess macrophage recruitment. qPCR was used to measure mRNA levels of proteins associated with a TAM phenotype as well as mRNA levels of MCP-1 and cyclooxygenase-2 (COX-2), an enzyme involved in PGE2 production. Results: Obese conditions resulted in increased macrophage recruitment and increased mRNA levels of TAM markers in macrophages. Furthermore, MCP-1 mRNA levels in adipose stromal cells and prostate cancer cells were higher under obese conditions compared to non-obese conditions. Similarly, obese conditions increased COX-2 mRNA levels in prostate cancer cells. Conclusions: Data from our in vitro study suggest that obesity may promote macrophage infiltration into the prostate tumor microenvironment and play a role in macrophage polarization towards a tumor-associated phenotype. This study will potentially serve to determine the value of macrophage and TAM concentration as biomarkers in this patient population, and lay the foundation for future studies investigating whether this phenomenon might be an effective therapeutic target. Citation Format: Gloria C. Galvan, Shruti Apte, Linda A. deGraffenried. Effects of obesity on regulation of macrophage population in the prostate tumor microenvironment. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A29.

Expression and significance of monocyte chemotactic protein 1 in early radiation-induced lung injury tissue of mice

Objective To study the expression and significance of monocyte chemotactic protein 1 (MCP-1) in the process of early radiation-induced lung injury in mice. Methods 40 mice were randomly divided into control group (10 cases) and post-irradiation group (30 cases). A large dose (15 Gy) irradiation on chest was performed in post-irradiation group. Every 10 mice in this group were killed on the 1st day (RT1d), the 10th day (RT10d), the 30th day (RT30d), respectively, and the specimens were collected. The expression of MCP-1 mRNA was detected by quantitative real-time PCR(RT-PCR), and MCP-1 and IL-6 proteins in BALF were detected by ELISA. The pathological changes of lung tissue were observed with HE staining. Results Under the microscope, lung tissue began to show inflammation on the 10th day and was more seriousness on the 30th day in post-irradiation group. Immunohistochemistry staining showed that the expression of MCP-1 in lung tissue irradiated was increased. RT-PCR results showed that the relative expressions of MCP-1 mRNA in control group, RT1d group, RT10d group and RT30d group were 7.63±0.77, 6.81±0.75, 5.75±0.73 and 4.61±0.75, respectively. The differences between RT1d group and control group, RT10d and RT1d group, and RT30d and RT10d were significantly different (all P < 0.05). ELISA results showed that the levels of MCP-1 and IL-6 proteins in post-irradiation group were significantly higher than those in the control group (P < 0.05). Conclusion The expression of MCP-1 in mice after irradiation is abnormal, and MCP-1 may participate in early radiation-induced lung injury. Key words: Radiation; Lung injury; Monocyte chemotactic protein 1; Interleukin-6

P65 down-regulates DEPTOR expression in response to LPS stimulation in hepatocytes

DEPTOR, a novel endogenous inhibitor of mTOR, plays an important role in regulating the inflammatory response in vascular endothelial cells (ECs) and in mouse skeletal muscle. However, the regulatory mechanism of DEPTOR transcription and its effects on liver inflammation are unknown presently. Here we reported the role of DEPTOR in regulating inflammatory response in mouse liver-derived Hepa1–6 cells and in a mouse model with LPS-induced hepatic inflammation. The results revealed that DEPTOR over-expression in Hepa1–6 liver cells increased the mRNA levels of the pro-inflammatory cytokines interleukin-6 (IL-6) and monocyte chemotactic protein-1 (MCP-1). Contrasting results were observed in Hepa1–6 cells with DEPTOR interference. Treatment Hepa1–6 cells with rapamycin, a specific inhibitor of mTORC1, increased MCP-1 mRNA, but have no significant effect on IL-6 mRNA. DEPTOR expression was down-regulated in Hepa1–6 cells with the treatment of inflammatory stimuli LPS or the over-expression of p65/NF-κB, a key inflammatory transcription factor. NF-κB antagonist (PDTC) and inhibitor (IκBα) blocked the effect of LPS on DEPTOR expression. The study in vivo showed that DEPTOR mRNA and protein were significantly reduced in a mouse model with LPS-induced hepatic inflammation, which was accompanied by a concurrent activation of the mTOR signaling pathway. Further, the transcriptional regulation of DEPTOR was explored, which revealed that DEPTOR promoter activity was significantly down-regulated by NF-κB. The progressive deletions and mutations demonstrated that the NF-κB binding motif situated at −145/−127 region is an essential component required for the DEPTOR promoter activity. Chromatin immunoprecipitation (ChIP) assays determined that p65 can directly interact with the DEPTOR promoter DNA. Those results indicate DEPTOR regulates liver inflammation at least partially via mTORC1 pathway, and is down-regulated by LPS through p65.

High glucose induced-macrophage activation through TGF-β-activated kinase 1 signaling pathway.

Transforming growth factor-β-activated kinase 1 (TAK1) plays a pivotal role in innate immune responses and kidney disease, and is critically involved in macrophage activation. However, there is a paucity of data to explore the role of high glucose (HG) in the regulation of TAK1 signaling and its functional role in macrophage activation. We assume that TAK1 signaling in hyperglycemic condition could be a key factor leading to macrophage activation and inflammation response. Mice macrophages were seeded on a 96-well cell culture plate; cell viability was tested after treatment with different concentration of TAK1 inhibitors. Cells were divided into groups (OZ300; MC; NC; HG; HG+OZ30, 100, 300nM) and treated for given time course. Monocyte chemotactic protein1(MCP-1) and tumor necrosis factor-α (TNF-α) mRNA levels were evaluated by qRT-PCR. Flow cytometry and confocal microscopy are used to analyse the activated macrophage induced by HG. Expression levels of p-TAK1, TAB 1, p-JNK, p-p38MAPK, NF-κBpp65 were detected by western blot. Nuclear translocation of NF-κBp65 was assessed by confocal microscopy. Our data revealed that high glucose not only significantly increased macrophage activation and subsequently abnormal high-expression of MCP-1 and TNF-α, but likewise remarkably enhanced TAK1 activation, MAPK phosphorylation, NF-κB expression in macrophages. Furthermore, pharmacological inhibition of TAK1 attenuated high glucose-triggered signal pathways, macrophage activation and inflammatory cytokines in a simulated diabetic environment. Our findings suggested that high glucose activated macrophages mainly in TAK1/MAPKs and TAK1/NF-κB-dependent manners, which lead to the polarization of macrophages towards a pro-inflammatory phenotype, and finally lead to diabetic nephropathy. In sum, the study raises novel data about the molecular mechanisms involved in the high glucose-mediated inflammatory response in macrophages.

The effect of necrostatin-1 on expression of liver monocyte chemotactic protein-1 in septic rats

To investigate the effect of necrostatin-1 (Nec-1) on the expression of liver monocyte chemotactic protein-1 (MCP-1) in septic rats and its mechanism. Forty-eight male Sprague-Dawley (SD) rats were randomly divided into sham group, model group, and Nec-1 group by randomized digital number method, with 16 rats in each group. The model of sepsis was reproduced by cecal ligation and puncture (CLP). Rats in sham group received anesthesia, and flipping the cecum followed by closure of the abdomen without ligation of the cecum. Rats in Nec-1 group were given 1 mg/kg Nec-1 [25 mg Nec-1 solution dissolved in 2.5 mL of dimethyl sulfoxide (DMSO)] through caudal vein 30 minutes before operation, while the rats in model group were given 0.1 mL/kg of DMSO only. Blood from abdominal aorta and liver tissue in each group were collected at 0 hour and 8 hours after operation. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were determined with automatic biochemistry analyzer. The pathological changes in liver were observed under light microscope using hematoxylin-eosin (HE) staining. The serum levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were determined by enzyme linked immunosorbent assay (ELISA). The MCP-1 mRNA expression in the liver was determined by reverse transcription-polymerase chain reaction (RT-PCR). There was no significant differences in the levels of serum ALT, AST, TNF-α, IL-6 and expressions of liver MCP-1 mRNA at 0 hour among three groups, and the liver cellular structure was normal. At 8 hours, compared with sham group, the expressions of serum ALT, AST, TNF-α, IL-6 and liver MCP-1 mRNA were significantly increased in model group and Nec-1 group [ALT (U/L): 172.35±21.88, 129.67±18.20 vs. 60.04±11.74, AST (U/L): 511.03±34.92, 363.51±25.25 vs. 254.83±31.04, TNF-α(ng/L): 603.96±24.18, 483.87±26.60 vs. 265.74±15.14, IL-6 (ng/L): 975.62±65.37, 712.09±45.47 vs. 310.42±13.88, MCP-1 mRNA (2-ΔΔCt): 7.09±0.18, 5.51±0.45 vs. 0.99±0.06, all P < 0.05]. Levels of the above parameters in Nec-1 group at 8 hours were significantly decreased compared with those of model group (all P < 0.05). Under light microscopy, it was noted that the structure of hepatic lobules was destroyed, with exacerbation of immunocyte infiltration at 8 hours in model group. At 8 hours, it was found that Nec-1 alleviated the pathological damage in Nec-1 group. Nec-1 can protect the liver of rats with sepsis, lower the expression of serum TNF-α and serum IL-6 and liver MCP-1 mRNA, and obviously reduce the damage of inflammation.

Forskolin Inhibits Lipopolysaccharide-Induced Modulation of MCP-1 and GPR120 in 3T3-L1 Adipocytes through an Inhibition of NFκB.

In an obese state, Toll-like receptor-4 (TLR-4) upregulates proinflammatory adipokines secretion including monocyte chemotactic protein-1 (MCP-1) in adipose tissue. In contrast, G-protein coupled receptor 120 (GPR120) mediates antiobesity effects. The aim of this study was to determine the signaling pathway by which Forskolin (FK), a cyclic adenosine monophosphate- (cAMP-) promoting agent causing positive changes in body composition in overweight and obese adult men, affects MCP-1 and GPR120 expression during an inflammatory response induced by lipopolysaccharide (LPS) in adipocytes, such as in an obese state. 3T3-L1 cells differentiated into adipocytes (DC) were stimulated with LPS in the absence or presence of FK and inhibitors of TLR-4 and inhibitor of kappa B (IκBα). In DC, LPS increased MCP-1, TLR-4, and nuclear factor-κB1 (NFκB1) mRNA levels, whereas it decreased GPR120 mRNA levels. In DC, FK inhibited the LPS-induced increase in MCP-1, TLR-4, and NFκB1 mRNA levels and the LPS-induced decrease in GPR120 mRNA. BAY11-7082 and CLI-095 abolished these LPS-induced effects. In conclusion, FK inhibits LPS-induced increase in MCP-1 mRNA levels and decrease in GPR120 mRNA levels in adipocytes and may be a potential treatment for inflammation in obesity. Furthermore, TLR-4-induced activation of NFκB may be involved in the LPS-induced regulation of these genes.

Chlorella Protein Hydrolysate Attenuates Glucose Metabolic Disorder and Fatty Liver in High-fat Diet-induced Obese Mice.

Chlorella (Parachlorella beijerinckii) powder is reported to show a preventive effect against metabolic syndromes such as arteriosclerosis, hyperlipidemia, and hypertension. Approximately 60% of the chlorella content is protein. In order to understand the role of chlorella protein, we prepared a chlorella protein hydrolysate (CPH) by protease treatment. Male C57BL/6 mice were divided into three groups: a normal diet group, high-fat diet (HFD) group, and high-fat diet supplemented with CPH (HFD+CPH) group. The CPH administration improved glucose intolerance, insulin sensitivity, and adipose tissue hypertrophy in the high-fat diet-fed mice. In addition, the HFD+CPH group had significantly decreased liver total cholesterol and triglyceride levels compared with those in the HFD group. Furthermore, the HFD+CPH group had a decreased level of monocyte chemotactic protein-1 (MCP-1) in serum and a lower MCP-1 mRNA expression level in adipose tissue compared with the HFD group. The present study suggests that chlorella protein hydrolysate can prevent a high-fat diet-induced glucose disorder and fatty liver by inhibiting adipocyte hypertrophy and reducing the MCP-1 protein and gene expression.

Effects of PB-EPCs on Homing Ability of Rabbit BMSCs via Endogenous SDF-1 and MCP-1

Traumas, infections, tumors, and some congenital malformations can lead to bone defects or even bone loss. The goal of the present study was to investigate whether inclusion of endothelial progenitor cells derived from peripheral blood (PB–EPCs) in cell-seeded partially deproteinized bone (PDPB) implants would stimulate recruitment of systemically injected bone marrow stromal cells (BMSCs) to the implant. Methods: BMSCs were injected intravenously with lentiviral expression vector expressing enhanced green fluorescent protein (eGFP) for tracing. Recruitment of eGFP-positive BMSCs was tested for the following implant configurations: 1) seeded with both BMSC and PB-EPC, 2) BMSC alone, 3) PB-EPC alone, and 4) unseeded PDPB. Protein and mRNA levels of endogenous stromal-derived factor-1 (SDF-1) and its receptor CXCR4, as well as monocyte chemotactic protein-1 (MCP-1) and its receptor CCR2, were evaluated on the 8th week. Immunohistochemical staining was performed to determine eGFP-positive areas at the defective sites. Masson’s trichrome staining was conducted to observe the distribution of collagen deposition and evaluate the extent of osteogenesis. Results: The mRNA and protein levels of SDF-1 and CXCR4 in the co-culture group were higher than those in other groups (p < 0.05) 8 weeks after the surgery. MCP-1 mRNA level in the co-culture group was also higher than that in the other groups (p < 0.05). Immunohistochemical assays revealed that the area covered by eGFP-positive cells was larger in the co-culture group than in the other groups (p < 0.05) after 4 weeks. Masson’s trichrome staining revealed better osteogenic potential of the co-culture group compared to the other groups (p < 0.05). Conclusion: These experiments demonstrate an association between PB-EPC and BMSC recruitment mediated by the SDF-1/CXCR4 axis that can enhance repair of bone defects.

Catechin-7-O-β-D-glucopyranoside isolated from the seed of Phaseolus calcaratus Roxburgh ameliorates experimental colitis in rats

The seeds of Phaseolus calcaratus Roxburgh (PHCR) are common legumes that comprise part of the daily diet in Chinese and Korean culture. Recent findings highlight anti-inflammatory and anti-septic potentials of catechin-7-O-β-D-glucopyranoside (CGP) isolated from PHCR seeds. We investigated the intestinal anti-inflammatory activity and associated mechanisms of CGP using a rat model of trinitrobenzenesulfonic acid (TNBS)-induced colitis. Oral treatment with CGP (10mg/kg body weight) suppressed body weight loss and intestinal inflammatory damages in TNBS-induced colitic rats. This treatment reduced myeloperoxidase activity and malondialdehyde level, but increased glutathione level in the TNBS colitic rats. CGP treatment also inhibited the TNBS-mediated increases in nitric oxide synthase, cyclooxygenase-2, interleukin-1β, tumor necrosis factor-α, intercellular adhesion molecule-1, and monocyte chemotactic protein-1 proteins or mRNA levels. This inhibition was accompanied by the increased mRNA levels of mucins MUC2 and MUC3. The CGP treatment prevented phosphorylation of p38 mitogen-activated protein kinase, IκB-α, and DNA-nuclear factor-κB binding, all of which were increased in the inflamed colons of TNBS-treated rats. Furthermore, oral administration with a crude PHCR butanol extract (100mg/kg body weight) which contains 1.5% of CGP showed intestinal anti-inflammatory potentials similar to that of CGP. Collectively, our current findings suggest that CGP or CGP-containing PHCR seeds may have favorable effects on intestinal inflammatory diseases.

Baicalein Inhibits Amadori-Glycated Albumin-Induced MCP-1 Expression in Retinal Ganglion Cells via a MicroRNA-124-Dependent Mechanism.

The purpose of this study was to characterize the inflammatory effect of amadori-glycated albumin (AGA) in cultured rat retinal ganglion cells (RGCs) and to further explore the potential mechanism of the anti-inflammatory effects of baicalein. Primary rat retinal neurons were separated and cultured. The levels of monocyte chemotactic protein-1 (MCP-1) mRNA and soluble MCP-1 produced by RGCs in response to AGA were measured with quantitative reverse transcription-PCR (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). In addition, the expression of microRNA-124 (miR-124) and histone deacetylases (HDACs) were detected in cultured rat RGCs by qRT-PCR in the presence or absence of baicalein. Luciferase reporter assays were used to validate the regulation of a putative target of miR-124. Amadori-glycated albumin stimulation increased the expression of MCP-1 and inhibited the expression of miR-124 in cultured rat RGCs. In addition, miR-124 directly controlled MCP-1 expression by binding directly to 3'-untranslated region (3'-UTR) of MCP-1. Next, we demonstrated that miR-124 expression was suppressed by HDACs and that treatment of RGCs with HDACs inhibitors increased miR-124 expression and decreased MCP-1 production. Furthermore, application of baicalein in RGCs attenuated AGA-induced MCP-1 expression and upregulated expression of miR-124 by controlling HDAC4 and HDAC5. Collectively, these data suggest that baicalein inhibits AGA-induced MCP-1 expression in retinal ganglion cells via a microRNA-124-dependent mechanism.

Prophylactic treatment with sulphonated immunoglobulin G attenuates development of mechanical allodynia-like response in mice with neuropathic pain.

Human immunoglobulin G (IgG) concentrates are immune-modulating, anti-inflammatory plasma-derived products. Clinical studies in recent years have suggested that IgG attenuates neuropathic pain. In this study, effects of sulphonated IgG on the development and maintenance of a mechanical allodynia-like response were examined in mice with neuropathic pain induced by a partial sciatic nerve ligation (PSL). When sulphonated IgG (400 or 1,000 mg/kg/day, i.p.) was administered for 5 days, from 1 day before surgery to post-operative day (POD) 3, the development of a mechanical allodynia-like response was attenuated. On the other hand, sulphonated IgG had little effect on the maintenance of a mechanical allodynia-like response when administered for 5 days, from POD 11 to POD 15, at which time a mechanical allodynia-like response had already been developed. To explore the mechanism of sulphonated IgG, the mRNA expression of inflammatory cytokines was evaluated in the injured sciatic nerve. Sulphonated IgG (1,000 mg/kg/day, i.p.) that was administered for 3 days, from 1 day before surgery to POD 1, significantly attenuated the up-regulation of tumor necrosis factor-α and monocyte chemotactic protein-1 mRNAs on POD 1. These results suggest that prophylactic treatment with sulphonated IgG attenuates the development of mechanical allodynia-like response by inhibition of inflammatory cytokine expression in mice with PSL.

Fucoxanthinol, Metabolite of Fucoxanthin, Improves Obesity-Induced Inflammation in Adipocyte Cells.

Fucoxanthin (Fx) is a marine carotenoid found in edible brown seaweeds. We previously reported that dietary Fx metabolite into fucoxanthinol (FxOH), attenuates the weight gain of white adipose tissue of diabetic/obese KK-Ay mice. In this study, to evaluate anti-diabetic effects of Fx, we investigated improving the effect of insulin resistance on the diabetic model of KK-Ay mice. Furthermore, preventing the effect of FxOH on low-grade chronic inflammation related to oxidative stress was evaluated on 3T3-L1 adipocyte cells and a RAW264.7 macrophage cell co-culture system. A diet containing 0.1% Fx was fed to diabetic model KK-Ay mice for three weeks, then glucose tolerance was observed. Fx diet significantly improved glucose tolerance compared with the control diet group. In in vitro studies, FxOH showed suppressed tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1) mRNA expression and protein levels in a co-culture of adipocyte and macrophage cells. These findings suggest that Fx ameliorates glucose tolerance in the diabetic model mice. Furthermore, FxOH, a metabolite of Fx, suppresses low-grade chronic inflammation in adipocyte cells.

Effects of intrathecal TRESK gene recombinant adenovirus on inflammatory responses mediated by chemokine in spinal cord of rats with neuropathic pain

Objective To evaluate the effects of intrathecal TRESK gene recombinant adenovirus on inflammatory responses mediated by chemokine in the spinal cord of rats with neuropathic pain (NP) . Methods Thirty-six male Sprague-Dawley rats, weighing 200-250 g, were randomly divided into 6 groups (n=6 each) using a random number table: control group (group C) ; sham operation group (group S) ; NP group; TRESK-overexpressed adenovirus group (group TRESK) ; negative adenovirus group (group Virus) ; normal saline group (group NS) . Spinal nerve injury was produced by exposing the sciatic nerve and its branches and ligation and transection of tibial nerve and common fibular nerve in anesthetized rats.In TRESK, Virus and NS groups, pAd/CMV/V5-DEST-TRESK 25 μl (109 IU/ml) , negative adenovirus 25 μl and normal saline 25 μl were intrathecally injected, respectively.At 1 day before operation (baseline, T0) and 1, 3, 7 and 14 days after operation (T1-4) , the mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency were measured.Six rats in each group were sacrificed after measurement of pain threshold at T3.The L4, 5 segments of the spinal cords were removed for determination of monocyte chemotactic protein-1 (MCP-1) , MIP-2, tumor necrosis factor-alpha (TNF-α) , interleukin-1 beta (IL-1β) and IL-6 mRNA expression by real-time PCR. Results There was no significant difference in thermal paw withdrawal latency at each time point between groups.Compared with C and S groups, MWT at T1-4 in NP and TRESK groups and at T1-3 in Virus and NS groups were significantly decreased, and the expression of MCP-1, MIP-2, TNF-α, IL-1β and IL-6 mRNA was up-regulated in NP, TRESK, Virus and NS groups.Compared with group NP, MWT was significantly increased at T1-4, and the expression of MCP-1, MIP-2, TNF-α, IL-1β and IL-6 mRNA was down-regulated in group TRESK. Conclusion The mechanism by which intrathecal TRESK gene recombinant adenovirus reduces NP is related to inhibition of inflammatory responses mediated by chemokine in the spinal cord of rats. Key words: Potassium channels; Neuralgia; Spinal cord; Receptors, CCR2; Chemokine CXCL2; Inflammatory

Sitagliptin Reduces Inflammation and Chronic Immune Cell Activation in HIV+ Adults With Impaired Glucose Tolerance.

HIV infection is associated with a greater risk for fasting hyperinsulinemia, impaired glucose tolerance, and higher incidence rates for vascular disease, myocardial infarction, or stroke despite effective combination antiretroviral therapy (cART). The underlying mechanism(s) may involve chronic low-grade systemic inflammation and immune cell activation. Dipeptidyl peptidase-4 inhibitors (sitagliptin) improve glucose tolerance and may possess immunomodulatory effects because leukocyte CD26 cell surface receptors express dipeptidyl peptidase-4 activity. Sitagliptin will reduce inflammatory and immune cell activation markers known to be elevated in cART-treated HIV-infected (HIV+) adults with impaired glucose tolerance. This was designed as a prospective, randomized, placebo-controlled, double-blind trial of sitagliptin in HIV+ adults. The setting was an academic medical center. Patients were cART-treated HIV+ men and women (n = 36) with stable HIV disease and impaired glucose tolerance. Interventions included sitagliptin 100 mg/d or placebo for 8 weeks. At baseline and week 8, plasma high-sensitivity C-reactive protein and C-X-C motif chemokine 10 concentrations (ELISA), oral glucose tolerance, and abdominal sc adipose mRNA expression for M1 macrophage markers (monocyte chemotactic protein-1, EGF-like module-containing, mucin-like hormone receptor 1). Sitagliptin reduced glucose area under the curve (P = .002) and improved oral glucose insulin sensitivity index (P = .04) more than placebo. Sitagliptin reduced plasma high-sensitivity C-reactive protein and C-X-C motif chemokine 10 levels more than placebo (P < .009). Adipose tissue monocyte chemotactic protein-1 mRNA abundance declined significantly more (P = .01), and adipose EGF-like module-containing, mucin-like hormone receptor 1 mRNA expression tended to decline more (P = .19) in sitagliptin than placebo. Sitagliptin had beneficial systemic and adipose anti-inflammatory effects in cART-treated HIV+ adults with impaired glucose tolerance. Large-scale, long-term studies should determine whether sitagliptin reduces cardiovascular risk and events in HIV+ adults.

Aromatase Controls Sjögren Syndrome–Like Lesions through Monocyte Chemotactic Protein-1 in Target Organ and Adipose Tissue–Associated Macrophages

Several autoimmune diseases are known to develop in postmenopausal women. However, the mechanism by which estrogen deficiency influences autoimmunity is unknown. Aromatase is an enzyme that converts androgens to estrogens. Herein, we used female aromatase gene knockout (ArKO) mice as a model of estrogen deficiency to investigate the molecular mechanism that underlies the onset and development of autoimmunity. Histological analyses showed that inflammatory lesions in the lacrimal and salivary glands of ArKO mice increased with age. Adoptive transfer of spleen cells or bone marrow cells from ArKO mice into recombination activating gene 2 knockout mice failed to induce the autoimmune lesions. Expression of mRNA encoding proinflammatory cytokines and monocyte chemotactic protein-1 increased in white adipose tissue of ArKO mice and was significantly higher than that in wild-type mice. Moreover, an increased number of inflammatory M1 macrophages was observed in white adipose tissue of ArKO mice. A significantly increased monocyte chemotactic protein-1 mRNA expression of the salivary gland tissue in ArKO was found together with adiposity. Furthermore, the autoimmune lesions in a murine model of Sjögren syndrome were exacerbated by administration of an aromatase inhibitor. These results suggest that aromatase may play a key role in the pathogenesis of Sjögren syndrome-like lesions by controlling the target organ and adipose tissue-associated macrophage.

MCP-1 as a potential target to inhibit the bone invasion by oral squamous cell carcinoma.

Bone invasion is a common complication of oral squamous cell carcinoma (OSCC), and this study sought to explore whether suppressed expression of monocyte chemotactic protein-1 (MCP-1) can be used to inhibit the bone invasion by OSCC. Strong staining of MCP-1 protein was observed from 10 archival blocks of OSCC by immunohistochemistry (IHC). Real-time PCR showed MCP-1 mRNA was highly expressed by OSCC cell lines (SCC25, HN5, and Tca8113), and SCC25 cells had the highest expression. An expression construct of a dominant negative variant of MCP-1 with 7 amino acids truncated (7ND), in the vector pcDNA was used to transfect SCC25 cells, and resultant stabilized SCC25 cells (SCC25-7ND) were generated by antibiotic selection. 10% conditioned media (CM, supernatant) of SCC25-7ND cells efficiently inhibited the formation of human osteoclasts grown from CD14(+) monocyte subpopulation, comparing with 10% CM of SCC25 cells. Further, cells of SCC25 or SCC25-7ND were injected onto the surface of calvariae of nude mice to establish an animal model of bone invasion by OSCC. H&E staining showed well-differentiated OSCC was formed in both groups, tumour cells invading the bone while osteoclasts locating in typical resorption lacunae. TRAP staining indicated significantly fewer osteoclasts were found in calvariae with cells of SCC25-7ND in comparison to cells of SCC25. These data demonstrate the relevance of MCP-1 with research on bone invasion by OSCC, and suggest the potential value of MCP-1 as a target to inhibit this common complication.

Activation of central muscarinic receptor type 1 prevents development of endotoxin tolerance in rat liver

Endotoxin tolerance is a mechanism in which cells receiving low doses of endotoxin, enter a transient phase with less inflammatory response to the next endotoxin challenges. Central nervous system is known to modulate systemic inflammation through activation of the cholinergic system; however, the role of central anti-inflammatory pathway in pathophysiology of hepatic endotoxin tolerance is unknown. Our study was designed to assess the effect central muscarinic type 1 receptor (M1) activation on development of endotoxin tolerance in rat liver. Endotoxin tolerance was induced by daily intraperitoneal injection of endotoxin (1mg/kg) for 5 days. Animals were randomly divided into two groups which received intracerebroventricular injection of either MCNA-343 (an M1 agonist, 5ng/kg) or saline 1h after intraperitoneal injection of saline or endotoxin. The responsiveness to endotoxin was assessed by measuring hepatic MCP-1 (monocyte chemotactic protein-1), iNOS (inducible nitric oxide synthase) and TNF-α (tumor necrosis-α) mRNA expression 3h after intraperitoneal administration of endotoxin using quantitative RT-PCR. A significant reduction in hepatic expression of MCP-1, iNOS and TNF-α was observed in rats with 5 days endotoxin challenge in comparison with rats given a single dose of endotoxin. There was no significant difference in hepatic expression of MCP-1, iNOS or TNF-α between acute and chronic LPS-treated groups in rats given MCNA-343. Central MCNA-343 stimulation could prevent the induction of hepatic endotoxin tolerance in animals receiving repeated doses of endotoxin. This indicates that M1 cholinergic receptor activation in the central nervous system can modulate endotoxin tolerance in rat liver.

Overexpression of juxtaposed with another zinc finger gene 1 reduces proinflammatory cytokine release via inhibition of stress‐activated protein kinases and nuclear factor‐κB

As an inhibitor of the nuclear receptor subfamily 2, group C, member 2 signaling pathway, juxtaposed with another zinc finger gene 1 (JAZF1) has been shown to be involved in gluconeogenesis, lipid metabolism, and insulin sensitivity. However, its role in hepatic lipogenesis and chronic low-grade inflammation leading to nonalcoholic fatty liver disease remains unknown. The aim of this study was to examine whether JAZF1 overexpression in vivo or in vitro can protect against palmitic acid (PA)-induced and high-fat diet (HFD)-induced systemic inflammatory responses, and the potential mechanism of this process. JAZF1 overexpression vector was transfected into PA-treated IAR-20 hepatocytes. The mRNA expression levels of proinflammatory cytokines were measured by real-time quantitative PCR, and stress-activated protein kinase activities were measured by immunoblotting. For in vivo studies, JAZF1 transgenic mice were fed an HFD for 12 weeks. Liver tissue was obtained for histological examination, real-time RT-PCR, and western blot analysis. PA significantly increased the expression levels of tumor necrosis factor-α, monocyte chemotactic protein-1 and interleukin-8 mRNA in IAR-20 hepatocytes in a dose-dependent and time-dependent manner. Treatment with JAZF1 or stress-activated protein kinase inhibitors inhibited PA-induced tumor necrosis factor-α, monocyte chemotactic protein-1 and interleukin-8 expression in these cells. In JAZF1-treated cells, the decreased expression of proinflammatory cytokines was accompanied by decreased p38 mitogen-activated protein kinase and c-Jun N-terminal kinase phosphorylation and increased nuclear factor-κB inhibitor-α protein levels, similarly to the role of signaling inhibitors. In vivo, HFD-induced expression of proinflammatory cytokines was markedly attenuated in JAZF1-Tg mice as compared with controls. This attenuation was accompanied by decreased activation of c-Jun N-terminal kinase, p38 mitogen-activated protein kinase, and nuclear factor-κB. These data provide evidence for the important role of JAZF1 in preventing lipogenesis and systemic inflammation-related disease.

Significant roles of the (pro)renin receptor in integrity of vascular smooth muscle cells

The (pro)renin receptor ((P)RR) is known to play an important role in the pathogenesis of vascular complications in diabetes mellitus and hypertension through its function in activating the local renin-angiotensin system. Recent studies have shown that the (P)RR is an accessory protein of the vacuolar H(+)-ATPase, suggesting a more fundamental and developmental function. In this study, smooth muscle cell-specific (P)RR/Atp6ap2 conditional knockout mice were generated. Smooth muscle cell-specific ablation of the (P)RR resulted in nonatherogenic sclerosis in the abdominal aorta. The deletion of the (P)RR did not affect ambulatory blood pressure levels. In cultured murine vascular smooth muscle cells (VSMCs), ablation of the (P)RR suppressed the expression of the Vo subunit c of the vacuolar H(+)-ATPase and impaired the cell recycling system, leading to autophagic cell death. In addition, loss of the (P)RR in VSMCs induced the expression of monocyte chemotactic protein-1 and interleukin-6 mRNAs. These results suggest that the (P)RR is essential for cell survival and downregulation of vascular inflammation in murine VSMCs through maintaining normal function of the vacuolar H(+)-ATPase.