Ursolic acid promotes robust tolerance to cardiac allografts in mice

Nuclear factor (NF)-κB is an important molecule in T cell activation. Our previous work has found that T cell-restricted NF-κB super-repressor (IκBαΔN-Tg) mice, expressing an inhibitor of NF-κB restricted to the T cell compartment, can permanently accept fully allogeneic cardiac grafts and secondary donor skin grafts. In this study, we explore if transient NF-κB inhibition by a small molecular inhibitor could induce permanent graft survival. Ursolic acid, a small molecular compound, dose-dependently inhibited T cell receptor (TCR)-triggered NF-κB nuclear translocation and T cell activation in vitro. In vivo, ursolic acid monotherapy prolonged significantly the survival of cardiac allograft in mice. Assisted with donor-specific transfusion (DST) on day 0, ursolic acid promoted 84·6% of first cardiac grafts to survive for more than 150 days. While the mice with long-term surviving grafts (LTS) did not reject the second donor strain hearts for more than 100 days without any treatment, they all promptly rejected the third-party strain hearts within 14 days. Interestingly, this protocol did not result in an increased proportion of CD4(+) CD25(+) forkhead box P3(+) regulatory T cells in splenocytes. That adoptive transfer experiments also did not support regulation was the main mechanism in this model. Splenocytes from LTS showed reduced alloreactivity to donor antigen. However, depletion of CD4(+) CD25(+) regulatory T cells did not alter the donor-reactivity of LTS splenocytes. These data suggest that depletion of donor-reactive T cells may play an important role in this protocol.     

Baicalein inhibits macrophage lipid accumulation and inflammatory response by activating the PPARγ/LXRα pathway

Lipid accumulation and inflammatory response are two major risk factors for atherosclerosis. Baicalein, a phenolic flavonoid widely used in East Asian countries, possesses a potential atheroprotective activity. However, the underlying mechanisms remain elusive. This study was performed to explore the impact of baicalein on lipid accumulation and inflammatory response in THP-1 macrophage-derived foam cells. Our results showed that baicalein up-regulated the expression of ATP binding cassette transporter A1 (ABCA1), ABCG1, liver X receptor α (LXRα), and peroxisome proliferator-activated receptor γ (PPARγ), promoted cholesterol efflux, and inhibited lipid accumulation. Administration of baicalein also reduced the expression and secretion of TNF-α, IL-1β, and IL-6. Knockdown of LXRα or PPARγ with siRNAs abrogated the effects of baicalein on ABCA1 and ABCG1 expression, cholesterol efflux, lipid accumulation as well as pro-inflammatory cytokine release. In summary, these findings suggest that baicalein exerts a beneficial effect on macrophage lipid accumulation and inflammatory response by activating the PPARγ/LXRα signaling pathway.     

Effects of FTO and PPARγ variants on intrauterine growth restriction in a Brazilian birth cohort

Intrauterine growth restriction (IUGR) is related to a higher risk of neonatal mortality, minor cognitive deficit, metabolic syndrome, and cardiovascular disease in adulthood. In previous studies, genetic variants in the FTO (fat mass and obesity-associated) and PPARγ (peroxisome proliferator-activated receptor-gamma) genes have been associated with metabolic disease, body mass index, and obesity among other outcomes. We studied the association of selected FTO (rs1421085, rs55682395, rs17817449, rs8043757, rs9926289, and rs9939609) and PPARγ (rs10865710, rs17036263, rs35206526, rs1801282, rs28763894, rs41516544, rs62243567, rs3856806, and rs1805151) single-nucleotide polymorphisms (SNPs) with IUGR, through a case-control study in a cohort of live births that occurred from June 1978 to May 1979 in a Brazilian city. We selected 280 IUGR cases and 256 controls for analysis. Logistic regression was used to jointly analyze the SNPs as well as factors such as maternal smoking, age, and schooling. We found that the PPARγ rs41516544 increased the risk of IUGR for male offspring (OR 27.83, 95%CI 3.65-212.32) as well as for female offspring (OR=8.94, 95%CI: 1.96-40.88). The FTO rs9939609 TA genotype resulted in a reduced susceptibility to IUGR for male offspring only (OR=0.47, 95%CI: 0.26-0.86). In conclusion, we demonstrated that PPARγ SNP had a positive effect and FTO SNP had a negative effect on IUGR occurrence, and these effects were gender-specific.     

Biochemical markers of bone metabolism in children with cow's milk allergy

Introduction

Patients with cow''s milk allergy (CMA) and following a cow milk protein-free diet for a long time are potentially at risk of developing bone abnormalities. To assess the balance between bone formation and resorption processes, we determined serum concentrations of osteocalcin (OC), bone alkaline phosphatase (BALP), C-terminal telopeptide of type I collagen (CTX), fetuin-A, osteoprotegerin (OPG) and receptor activator of nuclear factor κB ligand (RANKL) in children with CMA.

Material and methods

The study included 50 prepubertal children with diagnosed cow''s milk allergy, who were under systematic medical and nutritional care at the Institute of Mother and Child and 40 healthy counterparts as a control group. The concentrations of bone metabolism markers were determined by immunoenzymatic assays.

Results

The diets of all investigated children were correct in terms of phosphorus and magnesium contents but deficient in terms of calcium and vitamin D. Serum OC and CTX as well as fetuin-A concentrations were similar in both studied groups. The BALP activity was significantly (p < 0.05) higher in children with cow''s milk allergy than in the controls. Serum OPG concentration was comparable in both groups, but the RANKL level was higher (p < 0.05) in CMA children than in healthy ones. Hence, the ratio of OPG/RANKL was lower in children with CMA.

Conclusions

Our study demonstrates slight disturbances in the profile of bone metabolism markers in growing children with CMA. The increase in RANKL level and decrease in OPG/RANKL ratio may contribute to intensification of bone resorption in these patients.     

Roles of TNF-α, GSK-3β and RANKL in the occurrence and development of diabetic osteoporosis

Objective: To investigate the roles of TNF-α, GSK-3β and RANKL in the occurrence and development of diabetic osteoporosis. Methods: Diabetic rat model was established; tissue section technology was used to observe the situation of osteoporosis in diabetic rats; rat serum levels of OC, RANKL, GSK-3β, P38mapk, TNF-α and INS were detected by Elisa assay; osteoblasts and osteoclasts were primarily cultured and identified by immunohistochemistry and tartrate-resistant acid phosphatase (TRAP) staining respectively. The effects of GSK-3β inhibitors, lithium chloride, TNF-α antagonists and RANKL antagonists on the proliferation of osteoblasts and osteoclasts were evaluated; quantitative PCR was used to assess the effects of GSK-3β inhibitors, lithium chloride, on TNF-α and RANKL gene expression in osteoblasts and osteoclasts, and the effects of TNF-α and RANKL antagonists on GSK-3β gene expression in osteoblasts and osteoclasts. Results: Diabetic rat model was successfully established; osteoblasts and osteoclasts were successfully isolated and cultured. Elisa experiments showed that in diabetic model group, the levels of RANKL, GSK-3β, P38mapk and TNF-α were significantly increased, while the levels of osteocalcin (OC) and insulin (INS) were significantly reduced; MTT results showed that osteoclast proliferation in GSK-3β inhibitor and lithium chloride groups were weaker than the untreated group, while osteoclast proliferation in TNF-α antagonist group and RANKL antagonist Group was very close to the untreated group. Osteoblast proliferation in GSK-3β inhibitor and lithium chloride groups were weaker than the untreated group, while osteoblast proliferation in TNF-α antagonist group and RANKL antagonist group was higher than the untreated group. In all of the corresponding groups, cell proliferation in the diabetic group was stronger than the untreated group. In GSK-3β inhibitor and lithium oxide groups, TNF-α and RANKL gene expression levels were elevated, but TNF-α and RANKL gene expression levels in the diabetic group were slightly lower than the control group. GSK-3β gene expression level in TNF-α antagonist group and RANKL antagonist group was reduced; GSK-3β gene expression level in diabetic group was lower than the control group. Conclusion: In diabetic rats, TNF-α, GSK-3β and RANKL levels were elevated; GSK-3β could promote the proliferation of osteoblasts and osteoclasts, and inhibit the expression of TNF-α and RANKL; TNF-α and RANKL can suppress the proliferation of osteoblasts while had little effect on osteoclast proliferation; they also can promote the GSK-3β gene expression; interactions between the three broke the balance between osteoblasts and osteoclasts, leading to osteoporosis.     

Ginsenoside Rg1 suppressed inflammation and neuron apoptosis by activating PPARγ/HO-1 in hippocampus in rat model of cerebral ischemia-reperfusion injury

Generally accepted, inflammation and neuron apoptosis are two characterized pathological features of cerebral ischemia-reperfusion (IR) injury. Ginsenoside Rg1 was reported showing distinct neuroprotective effect in cerebral IR injury but the underlying mechanisms are still unclear. PPARγ/Heme oxygenase-1 (HO-1) signaling was proved effective in suppressing both apoptosis and inflammation. This study was aimed to investigate whether PPARγ/HO-1 signaling was involved in cerebral IR injury and ginsenoside Rg1’s neuroprotective effect in cerebral IR injury. Cerebral IR injury was induced by middle cerebral artery occlusion in rats. The PPARγ agonist rosiglitazone (ROZ) and the HO-1 inhibitor zinc protoporphyrin-IX (ZnPP) and ginsenoside Rg1 at various concentrations were used to treat the modeled rats. Neurological deficits, apoptosis and inflammation in hippocampus were evaluated. Furthermore, HO-1 enzymatic activity, expression levels of apoptosis-related and inflammation-related proteins, concentrations of inflammatory cytokines were also determined. The results showed that PPARγ activation by ROZ significantly attenuated neurological deficits, apoptosis and inflammation in hippocampus in cerebral IR rats. However, the neuroprotective effect of ROZ was then impaired by HO-1 inhibitor ZnPP. This effect was evidenced by changes of expression levels of PPARγ, bcl-2, cleaved caspase-3, cleaved caspase-9, IL-1β, TNF-α, HMGB1, and RAGE in hippocampus of modeled animals. Ginsenoside Rg1 showed similar effect to ROZ in activating PPARγ/HO-1 in protecting against apoptosis and inflammation but also impaired by ZnPP administration. In conclusion, PPARγ/HO-1 signaling was critical in mediating apoptosis and inflammation, which was also the therapeutic target of ginsenoside Rg1 in cerebral IR injury.     

Antidiabetic effects of Corni Fructus extract in streptozotocin-induced diabetic rats

Purpose

Diabetes is the leading cause of end-stage renal failure. The present study was undertaken to characterize the effects of Corni Fructus on diabetic nephropathy in streptozotocin-induced diabetic rats and their mechanisms.

Materials and Methods

Streptozotocin-diabetic rats were orally administrated with Corni Fructus at a dose of 100, 200 or 400 mg/kg body mass for 40 days.

Results

Corni Fructus-treated diabetic rats showed significant decreases of blood glucose, urinary protein levels and water consumption. Corni Fructus also reduced serum total cholesterol, total triglyceride and low-density lipoprotein cholesterol levels, and showed a tendency of enhancing high-density lipoprotein cholesterol level. Levels of serum albumin and creatinine in diabetic rats were also significantly reduced by Corni Fructus administration at a dose of 200 and 400 mg/kg body mass compared with non-treated diabetic rats. Corni Fructus increased catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidose (GSH-px) activities in the kidneys of diabetic rats. Furthermore, Corni Fructus treatment enhanced renal peroxisome proliferator-activated receptor-γ (PPARγ) expression in diabetic rats.

Conclusion

These results demonstrated that Corni Fructus may have the potential to protect the animals from diabetic nephropathy by amelioration of oxidative stress and stimulation of PPARγ expression.     

Genistein attenuates glucocorticoid-induced bone deleterious effects through regulation Eph/ephrin expression in aged mice

Objective: This study was performed to investigate bone deteriorations and the involvement of skeletal Eph/ephrin signaling pathway of GIOP aged mice in response to the treatment of genistein. Methods: The biomarkers in serum and urine were measured, tibias were taken for the measurement on gene and protein expression and histomorphology analysis, and femurs were taken for the measurement on bone Ca and three-dimensional architecture of trabecular bone. Results: Genistein showed a greater increase in bone Ca, BMD and significantly increased FGF-23 and OCN, reduced TRACP-5b, PTH and CTX in GIOP mice. Genistein reversed DXM-induced trabecular deleterious effects and stimulated bone remodeling. The treatment of DXM group with genistein significantly elevated the ratio of OPG/RANKL. Moreover, genistein administration down-regulated the mRNA and protein expression of Eph A2 and ephrin A2 in tibia of the GIOP mice. In contrast, the mRNA and protein expression of Eph B4 and ephrin B2 were increased in mice treated by DXM with genistein as compared to the DXM single treatment. Conclusions: DXM-induced trabecular bone micro-structure deterioration in aged mice was involved in the regulation of the Eph receptors and ephrin ligands. Genistein might represent a therapy with bone-forming as well as an anti-resorptive activity in GIOP mice. The underlying mechanism was mediated, at least partially, through regulation Eph/ephrin signaling.     

Mannose prevents acute lung injury through mannose receptor pathway and contributes to regulate PPARγ and TGF-β1 level

Mannose has been reported to prevent acute lung injury (ALI), and mannose receptor (MR) has been demonstrated to have a role. The rationale for this study is to characterize the mechanism by which mannose and MR prevent lipopolysaccharide (LPS)-induced ALI. Male ICR mice were pretreated mannose by intravenous injection 5 min before and 3 h after intratracheal instillation of LPS. Pathological changes, proinflammatory mediator, peroxisome proliferator activated receptor gamma (PPARγ), MR, and transforming growth factor β1 (TGF-β1) levels were determined. The RAW264.7 cells were pretreated with mannose and stimulated with LPS for 3 h. Proinflammatory mediator and TGF-β1 in the culture media, PPARγ, MR, and TGF-β1 expression in RAW 264.7 cells were measured. Mannose markedly attenuated the LPS-induced histological alterations and inhibited the production of proinflammatory mediator in mice and in RAW 264.7 cells. Mannose increased PPARγ and MR expression, and inhibited TGF-β1 stimulated by LPS. Interestingly, competitive inhibition of MR with mannan was associated with elimination of the anti-inflammatory effects of mannose, and reversed effects of mannose of regulation to PPARγ and TGF-β1. MR is important in increasing PPARγ and decreasing TGF-β1 expression and plays a critical role in mannose’s protection against ALI.     

Bacterial extract OM-85 BV protects mice against experimental chronic rhinosinusitis

Objectives: To investigate the therapeutic effects of OM-85 BV as an adjunctive treatment on experimental chronic rhinosinusitis (CRS) in mice. Methodology: Female BALB/c mice aged 8-12 weeks were sensitized and administrated by intranasal Aspergillus fumigatis (AF) three times per week for 1 week, 3 weeks, 2 months and 3 months (n = 10 each time point). The mice were randomly and equally assigned to four groups: normal control group, model group, OM-85-BV plus amoxicillin group, and isolated amoxicillin group. Inflammatory changes were determined by hematoxylin-eosin (HE) staining. The expression levels of suppressor of cytokine signaling (SOCS) 1, SOCS3, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ in samples were assessed by using real-time PCR (RT-PCR) and Western blotting. Results: There were significantly inflammatory and structural changes between the model and other groups. Compared to the model group, the mRNA expression levels of SOCS1, SOCS3, TNF-α, and IFN-γ were significantly decreased in OM-85-BV plus amoxicillin group and isolated amoxicillin group, along with the protein levels. Conclusion: The bacterial extract OM-85 BV is a low-cost alternatively adjunctive drug to treat CRS with simple oral administration, good safety, and few side effects.     

Thrap3 docks on phosphoserine 273 of PPARγ and controls diabetic gene programming

Phosphorylation of peroxisome proliferator-activated receptor γ (PPARγ) at Ser273 by cyclin-dependent kinase 5 (CDK5) in adipose tissue stimulates insulin resistance, but the underlying molecular mechanisms are unclear. We show here that Thrap3 (thyroid hormone receptor-associated protein 3) can directly interact with PPARγ when it is phosphorylated at Ser273, and this interaction controls the diabetic gene programming mediated by the phosphorylation of PPARγ. Knockdown of Thrap3 restores most of the genes dysregulated by CDK5 action on PPARγ in cultured adipocytes. Importantly, reduced expression of Thrap3 in fat tissue by antisense oligonucleotides (ASOs) regulates a specific set of genes, including the key adipokines adiponectin and adipsin, and effectively improves hyperglycemia and insulin resistance in high-fat-fed mice without affecting body weight. These data indicate that Thrap3 plays a crucial role in controlling diabetic gene programming and may provide opportunities for the development of new therapeutics for obesity and type 2 diabetes.     

Sodium fusidate ameliorates the course of diabetes induced in mice by multiple low doses of streptozotocin

We studied the effects of the immunosuppressant sodium fusidate (fusidin) on murine immunoinflammatory diabetes mellitus (DM) induced by multiple low doses of streptozotocin (SZ). Fusidin was given by gavage to three strains of mice (C57KsJ, C57BL/6, CD1) at doses 10 or 100 mg/kg body weight every other day. The drug was administered as an early or late prophylactic regime starting either 1 day prior to the first or after the fifth and last injection of SZ. In both situations the largest dose of fusidin successfully reduced the clinical, chemical and histological signs of DM, the treated mice having significantly lower glycaemic values and milder (often absent) insulitis compared with sham-treated animals or controls given SZ alone. The antidiabetogenic effect was long-lasting as it was maintained up to 1 month after cessation of therapy. In contrast, fusidin prophylaxis failed to prevent development of hyperglycaemia acutely induced by one single and high (160 mg/kg) dose of SZ, which is a model of DM primarily due to the toxic action of SZ on the beta cells and does not involve immunopathogenetic mechanisms.On day 14 after SZ, fusidin markedly altered the circulating cytokine profile induced in vivo by ConA, reducing the levels of IFN-gamma, IL-2 and TNF-alpha and augmenting the level of IL-6. However, only the inhibitory effect of the drug on the synthesis/release of IFN-gamma seemed to be causally related to its capacity to counteract the SZ-induced DM. In fact, the disease was prevented by a neutralizing monoclonal antibody (mAb) against IFN-gamma, but not by anti-IL-2 receptor mAb, a soluble form of TNF-receptor type 1 or recombinant human IL-6. The prevention of disease by fusidin was also partly reversed by exogenously administered recombinant mouse IFN-gamma.The data provide further in-vivo evidence for the anti-diabetogenic and immunomodulatory properties of fusidin and indicate that this drug could have a role in prevention and treatment of human type 1 DM.     

TNFalpha inhibition in MRL/lpr mice ameliorates pulmonary but not renal disease

TNFalpha inhibition has a clearly beneficial effect in a number of arthritides and in Crohn's disease. The exact mechanism of action is uncertain with studies showing inhibition of chemokines, inhibition of adhesion molecule expression, and improved T-cell function. Unlike most therapeutic interventions for autoimmune disease, TNFalpha inhibition appears to act on specific pathologic processes. It is not known how wide-spread these TNFalpha-mediated pathologic processes are. Efforts to expand the use of TNFalpha inhibition have had notable successes but have been disappointing in other disorders. We hypothesized that TNFalpha-mediated pathologic processes might play a significant role in the end-organ effects seen in SLE. We modeled SLE by using MRL/lpr mice and treated with two types of TNFalpha inhibitor. Pulmonary disease was significantly improved in the treated groups compared to controls. In contrast, renal disease was unaffected suggesting that in lupus, where multiple organs are affected, different pathologic processes may be mediating the end-organ damage. This has important implications for designing therapeutics for SLE.     

The effects of hepatic ischemia/reperfusion injury on postoperative cognitive function in aged rats

IntroductionHepatic ischemia/reperfusion injury (I/R) is a significant source of morbidity and mortality after liver surgery. The aim of this study was to investigate the effect of hepatic I/R injury on the hippocampus in rats with postoperative cognitive dysfunction (POCD).Material and methodsAdult male Sprague-Dawley rats (n = 160, age: 20–25 months, weight: 300–350 g) received I/R surgery with ischemia for 20 min, 30 min, and 40 min in different groups. Behavior tests of the Morris water maze (MWM) test and the passive avoidance test were applied. Population spike (PS) of pyramidal cells, nuclear factor κB (NF-κB) and protein kinase γ (PKCγ) in the hippocampus were observed.ResultsWithin 10 days after surgery, in aged rats with varying impaired cognitive function, spike size and spike latency period were reduced, level of PKCγ was decreased and an increased level of NF-κB was observed in the I/R group, especially in the I/R group with ischemia for 40 min. The parameters showed no significant difference in rats in the I/R group compared with the sham group at the 18^th day after surgery.ConclusionsHepatic I/R injury has a negative impact on the postoperative cognitive function in aged rats, leading to hippocampus changes with PS abnormity and level changes of NF-κB, PKCγ. However, this cognitive deficit improved over time.