Chung-Yuh Tzeng

Leptin induces IL-8 expression via leptin receptor, IRS-1, PI3K, Akt cascade and promotion of NF-κB/p300 binding in human synovial fibroblasts

Leptin, the adipocyte-secreted hormone that centrally regulates weight control, is known to function as an immunomodulatory regulator. We investigated the signaling pathway involved in IL-8 production caused by leptin in both rheumatoid arthritis synovial fibroblasts (RASF) and osteoarthritis synovial fibroblasts (OASF). RASF and OASF expressed the long (OBRl) and short (OBRs) isoforms of the leptin receptor. Leptin caused concentration- and time-dependent increases in IL-8 production. Leptin-mediated IL-8 production was attenuated by OBRl receptor antisense oligonucleotide, JAK2 inhibitor or STAT3 small interference RNA (siRNA). Transfection with insulin receptor substrate (IRS)-1 siRNA or dominant-negative mutant of p85 and Akt or pretreatment with phosphatidylinositol 3-kinase inhibitor (Ly294002 and wortmannin), Akt inhibitor, NF-kappaB inhibitor (PDTC) and NF-kappaB inhibitor peptide also inhibited the potentiating action of leptin. Stimulation of RASF with leptin activated IkappaB kinase alpha/beta (IKK alpha/beta), p65 phosphorylation at Ser(276), p65 translocation from the cytosol to the nucleus, and kappaB-luciferase activity. Moreover, pretreatment with p300 inhibitor (curcumin) also blocked IL-8 expression. The binding of p65 to the NF-kappaB elements, as well as the recruitment of p300 and the enhancement of histone H3 acetylation on the IL-8 promoter was enhanced by leptin, which was inhibited by wortmannin, Akt inhibitor or IRS-1 siRNA. These results suggest that leptin increased IL-8 production in synovial fibroblast via the OBRl/JAK2/STAT3 pathway, as well as the activation of IRS1/PI3K/Akt/NF-kappaB-dependent pathway and the subsequent recruitment of p300.

Peptidoglycan Enhances IL-6 Production in Human Synovial Fibroblasts via TLR2 Receptor, Focal Adhesion Kinase, Akt, and AP-1- Dependent Pathway

Peptidoglycan (PGN), the major component of the cell wall of Gram-positive bacteria, activates the innate immune system of the host and induces the release of cytokines and chemokines. We investigated the signaling pathway involved in IL-6 production stimulated by PGN in rheumatoid arthritis synovial fibroblasts. PGN caused concentration- and time-dependent increases in IL-6 production. PGN-mediated IL-6 production was attenuated by TLR2 small interfering RNA and nucleotide-binding oligomerization domain 2 small interfering RNA. Pretreatment with PI3K inhibitor (Ly294002 and wortmannin), Akt inhibitor, and AP-1 inhibitor (tanshinone IIA) also inhibited the potentiating action of PGN. PGN increased the focal adhesion kinase (FAK), PI3K, and Akt phosphorylation. Stimulation of rheumatoid arthritis synovial fibroblast cells with PGN increased the accumulation of phosphorylated c-Jun in the nucleus, AP-1-luciferase activity, and c-Jun binding to the AP-1 element on the IL-6 promoter. PGN mediated an increase in the accumulation of phosphorylated c-Jun in the nucleus, AP-1-luciferase activity, and c-Jun binding to AP-1 element was inhibited by Ly294002, Akt inhibitor, and FAK mutant. Our results suggest that PGN increased IL-6 production in human synovial fibroblasts via the TLR2 receptor/FAK/PI3K/Akt and AP-1 signaling pathway.

Adiponectin increases MMP‐3 expression in human chondrocytes through adipor1 signaling pathway

Articular adipose tissue is a ubiquitous component of human joints, and adiponectin is a protein hormone secreted predominantly by differentiated adipocytes and involved in energy homeostasis. The adiponectin is significantly higher in synovial fluid of patients with osteoarthritis and rheumatoid arthritis. Matrix metalloproteinases (MMP)‐3 may contribute to the breakdown of articular cartilage during arthritis. We investigated the signaling pathway involved in MMP‐3 caused by adiponectin in human chondrocytes. Adiponectin increased the secretion of MMP‐3 in cultured human chondrocytes, as shown by qPCR, Western blot, and ELISA analysis. Adiponectin‐mediated MMP‐3 expression was attenuated by AdipoR1 but not AdipoR2 siRNA. Pretreatment with 5′‐AMP‐activated protein kinase (AMPK) inhibitor (araA and compound C), p38 inhibitor (SB203580), and NF‐κB inhibitor (PDTC and TPCK) also inhibited the potentiating action of adiponectin. Activations of p38, AMPK, and NF‐κB pathways after adiponectin treatment were demonstrated. Taken together, our results provide evidence that adiponectin acts through AdipoR1 to activate p38 and AMPK, resulting in the activations of NF‐κB on the MMP‐3 promoter and contribute cartilage destruction during arthritis. J. Cell. Biochem. 112: 1431–1440, 2011.

Acute effect of electroacupuncture at the Zusanli acupoints on decreasing insulin resistance as shown by lowering plasma free fatty acid levels in steroid-background male rats

BackgroundInsulin sensitivity has been enhanced by electroacupuncture (EA) in rats, but the EA phenomenon in an insulin resistant state is still unclear. This study reports the use of a large dose of prednisolone to evaluate the effects of EA in a state of insulin resistance.MethodsThe plasma levels of free fatty acids (FFAs) were estimated in steroid-background rats (SBRs) and compared with those in healthy rats treated with normal saline. In addition, plasma glucose and endogenous insulin levels were assayed to calculate the homeostasis model assessment (HOMA) index. Intravenous glucose tolerance test (IVGTT) was carried out to compare glucose tolerance. The SBRs were randomly divided into EA-treatment and non-EA treatment groups and 15-Hz EA was applied to the bilateral Zusanli acupoints to investigate its effects on insulin resistance. In addition to an insulin challenge test (ICT) and IVGTT, the plasma levels of FFAs were measured and western blot was performed to help determine the effects of EA on the insulin resistant state.ResultsThe plasma levels of FFAs increased markedly in SBRs, the HOMA index was markedly higher, and glucose tolerance was impaired. EA improved glucose tolerance and insulin sensitivity by decreasing the plasma levels of FFAs. Further, the insulin signaling proteins (IRS1) and glucose transporter isoform protein (GLUT4) in skeletal muscle inhibited by prednisolone recovered after EA.ConclusionInsulin resistance was successfully induced by a large dose of prednisolone in male rats. This insulin resistance can be improved by 15 Hz EA at the bilateral Zusanli acupoints, as shown by decreased plasma levels of FFAs.

Bradykinin-induced IL-6 expression through bradykinin B2 receptor, phospholipase C, protein kinase Cδ and NF-κB pathway in human synovial fibroblasts

Bradykinin (BK) is an inflammatory mediator, and shows elevated levels in regions of severe injury and inflammatory diseases. It has been shown to induce interleukin-6 (IL-6) expression in inflammatory responses in rheumatoid arthritis. We investigated the signaling pathway involved in IL-6 production caused by BK in synovial fibroblasts. BK caused concentration- and time-dependent increases in IL-6 production. By using pharmacological inhibitors or genetic inhibition of the BK receptor, siRNA revealed that B2 but not B1 BK receptors are involved in BK-mediated up-regulation of IL-6. BK-mediated IL-6 production was attenuated by phospholipase C inhibitor (U73122), protein kinase Cdelta inhibitor (rottlerin), NF-kappaB inhibitor (PDTC), IkappaB protease inhibitor (TPCK) and NF-kappaB inhibitor peptide. Stimulation of synovial fibroblasts with BK activated IkappaB kinase alpha/beta (IKK alpha/beta), IkappaBalpha phosphorylation, IkappaBalpha degradation, p65 phosphorylation at Ser(276), p65 and p50 translocation from the cytosol to the nucleus and kappaB-luciferase activity. BK mediated an increase of IKK alpha/beta and IkappaBalpha phosphorylation, kappaB-luciferase activity and p65 and p50 binding to the NF-kappaB element was inhibited by B2 BK receptor antagonist (HOE140), U73122 and rottlerin. Our results suggest that BK increased IL-6 production in synovial fibroblasts via the B2 BK receptor/PI-PLC/PKCdelta/and NF-kappaB signaling pathway.

Electroacupuncture at the Zusanli (ST-36) Acupoint Induces a Hypoglycemic Effect by Stimulating the Cholinergic Nerve in a Rat Model of Streptozotocine-Induced Insulin-Dependent Diabetes Mellitus

Animal studies have shown that electroacupuncture (EA) at Zusanli (ST-36) and Zhongwan (CV-12) acupoints reduces plasma glucose concentrations in rats with type II diabetes. However, whether EA reduces plasma glucose levels in type I diabetes is still unknown. In this study, we explore the various non-insulin-dependent pathways involved in EA-induced lowering of plasma glucose. Streptozotocin (STZ) (60 mg kg−1, i.v.) was administered via the femoral vein to induce insulin-dependent diabetes in non-adrenalectomized and in adrenalectomomized rats. EA (15 Hz) was applied for 30 min to bilateral ST-36 acupoints after administration of Atropine (0.1 mg kg−1 i.p.), Eserine (0.01 mg kg−1 i.p.), or Hemicholinium-3 (5 μg kg−1 i.p.) in non-adrenalectomized rats. Rats administered acetylcholine (0.01 mg kg−1 i.v.) did not undergo EA. Adrenalectomized rats underwent EA at bilateral ST-36 acupoints without further treatment. Blood samples were drawn from all rats before and after EA to measure changes in plasma glucose levels. Expression of insulin signaling proteins (IRS1, AKT2) in atropine-exposed rats before and after EA was measured by western blot. Atropine and hemicholinium-3 completely blocked the plasma glucose lowering effects of EA, whereas eserine led to a significant hypoglycemic response. In addition, plasma glucose levels after administration of acetylcholine were significantly lower than the fasting glucose levels. In STZ-adrenalectomized rats, EA did not induce a hypoglycemic response. EA stimulated the expression of IRS1 and AKT2 and atropine treatment blocked the EA-induced expression of those insulin signaling proteins. Taken together, EA at the ST-36 acupoint reduces plasma glucose concentrations by stimulating the cholinergic nerves.

A combined therapy using stimulating auricular acupoints enhances lower-level atropine eyedrops when used for myopia control in school-aged children evaluated by a pilot randomized controlled clinical trial

OBJECTIVE This study was designed to compare the reduction in myopia progression in patients treated with atropine eyedrops alone with patients treated with a combined treatment of atropine and stimulation of the auricular acupoints. METHODS This study was a randomized single-blind clinical controlled trial. A total of 71 school-aged children with myopia, who fulfilled the eligibility criteria, were recruited. They were randomly assigned into three groups. These were 22 treated with the 0.25% atropine (0.25A) only, 23 treated with the 0.5% atropine (0.5A) only and 26 treated with 0.25% atropine together with stimulation of the auricular acupoints (0.25A+E). The differences in the post-treatment effects among these three groups were statistically assessed. The primary outcome parameter was myopia progression, which was defined as diopter change per year (D/Y) after cycloplegic refraction measurement. RESULTS The mean myopia progression of the 0.25A group was 0.38+/-0.32 D/Y. No significant difference in mean myopia progression was found between the 0.5A (0.15+/-0.15 D/Y) and 0.25A+E (0.21+/-0.23 D/Y) groups. However, there was a markedly reduced myopia progression in the 0.25A+E group compared to the 0.25A group (p<0.05). Furthermore, there was no statistical difference among these three groups in axial length elongation (ALE) of eye during this stage of the investigation. CONCLUSIONS This study demonstrates that there was efficacy in stimulating the auricular acupoints and this enhanced the action of 0.25% atropine as a means of myopia control. The result was an effect almost equal to that of 0.5% atropine alone. There is also a need that the ALE of the eye should be further investigated over a longer period using the combined therapy.

SUPERFICIAL RADIAL NERVE COMPRESSION CAUSED BY A PAROSTEAL LIPOMA OF PROXIMAL RADIUS: A CASE REPORT

The superficial radial nerve might be compressed or injured at various anatomical sites along its course in the forearm. Most of the superficial radial nerve neuropathy are caused by pathological lesions such as trauma, a mass or tight band at the distal third of the forearm. Wartenbergs syndrome is the most common cause of sensory radial entrapment at the distal forearm. Compression of superficial radial nerve occurring at the proximal third of forearm is unusual. We present a rare case of superficial radial nerve compression due to a parosteal lipoma of proximal radius. Results of complete physical and radiological examinations are also presented. Surgical intervention of the tumour mass was performed for nerve decompression. The patient reported total relief of the neurological symptom post-operatively. This rare case demonstrates the unique characteristics of parosteal lipoma with unusual superficial radial nerve neuropathy at the proximal radius. This report reminds us that there is the possibility of superficial nerve compression caused by tumour mass over the proximal third of forearm.

Electroacupuncture improves glucose tolerance through cholinergic nerve and nitric oxide synthase effects in rats

The purpose of this investigation was to evaluate the effect and mechanisms of electroacupuncture (EA) at the bilateral Zusanli acupoints (ST-36) on glucose tolerance in normal rats. Intravenous glucose tolerance test (IVGTT) was performed to examine the effects of electroacupuncture (EA) on glucose tolerance in rats. The EA group underwent EA at the ST-36, with settings of 15 Hz, 10 mA, and 60 min; the control group underwent the same treatments, but without EA. Atropine, hemicholinium-3 (HC-3) or NG-nitro-L-arginine methyl ester (L-NAME) were injected into the rats alone or simultaneously and EA was performed to investigate differences in plasma glucose levels compared to the control group. Plasma samples were obtained for assaying plasma glucose and free fatty acid (FFA) levels. Western blot was done to determine the insulin signal protein and nNOS to exam the correlation between EA and improvement in glucose tolerance. The EA group had significantly lower plasma glucose levels compared to the control group. Plasma glucose levels differed significantly between the EA and control groups after the administration of L-NAME, atropine, or HC-3 treatments alone, but there were no significant differences in plasma glucose with combined treatment of L-NAME and atropine or L-NAME and HC-3. EA decreased FFA levels and enhanced insulin signal protein (IRS1) and nNOS activities in skeletal muscle during IVGTT. In summary, EA stimulated cholinergic nerves and nitric oxide synthase for lowering plasma FFA levels to improve glucose tolerance.

Differential expression profiling of orange-spotted grouper larvae, Epinephelus coioides (Hamilton), that survived a betanodavirus outbreak.

Nervous necrosis virus (NNV), a piscine nodavirus, has caused serious viral nervous necrosis and viral encephalopathy and retinopathy in hatchery-reared larvae and juveniles of a wide range of marine teleost species worldwide in the last two decades. Although the mortality of NNV-infected larvae is nearly 100%, there are still some larvae that survive this catastrophe. To comprehensively understand the variations of these survivors at the molecular level, we collected orange-spotted grouper larvae that survived an NNV outbreak in an indoor hatchery in southern Taiwan to study differential gene expression. Healthy larvae with high, medium and low levels of detected NNV were compared with morbid larvae using a 9600-clone-containing grouper larva cDNA microarray, and differential gene expression was further confirmed by a quantitative real-time polymerase chain reaction. Significant variation exists in healthy larvae. The following genes were upregulated: adenylate kinase 1-2, myosin binding protein H-like, myosin light chain 2, myosin light chain 3, tropomyosin, fast/white muscle troponin T embryonic isoform, and parvalbumin 1 and 2 genes. The following genes were downregulated: apolipoprotein A-I, trypsinogen, pyruvate kinase and astacin-like metalloprotease. Moreover, immunoglobulin M heavy chain gene transcription was significantly higher in healthy larvae that had high virus levels, indicating that humoral immunity might protect organisms from viral infection. These results suggest that some non-immune-related genes may have played important roles in survival during the larval metamorphosis stage, after betanodavirus infection.