Assessment of the antimicrobial and immunomodulatory activity of QS-CATH, a promising therapeutic agent isolated from the Chinese spiny frogs (Quasipaa spinosa).

Cathelicidins are important antimicrobial peptides in various vertebrate species where they are crucial parts of the innate immune system. The current understanding of amphibian cathelicidins is limited, particularly with regard to their immunomodulatory effects. To address this knowledge gap, we produced the cDNA sequence of the cathelicidin gene from a skin transcriptome of the Chinese spiny frog Quasipaa spinosa. The amino acid sequence of the Quasipaa spinosa cathelicidin (QS-CATH) was predicted to consist of a signal peptide, a cathelin domain, and a mature peptide. Comparative analysis of the QS-CATH amino acid sequence with that of other amphibian cathelicidins revealed high variability in the functional mature peptide among amphibians, whereas the cathelin domain was conserved. The QS-CATH gene was expressed in several tissues, with the highest level of expression in the spleen. Upregulation of QS-CATH after Aeromonas hydrophila infection occurred in the kidney, gut, spleen, skin, and liver. Chemically synthesized QS-CATH exhibited pronounced antibacterial activity against Shigella flexneri, Staphylococcus warneri, Escherichia coli, Salmonella enterica, and Listeria monocytogenes. Furthermore, QS-CATH disrupted the cell membrane integrity of S. flexneri, as evidenced by a lactate dehydrogenase release assay, and it hydrolyzed the genomic DNA of S. flexneri. Additionally, QS-CATH elicited chemotaxis and modulated the expression of inflammatory cytokine genes in RAW264.7 mouse leukemic monocyte/macrophage cells. These findings confirm the antimicrobial effects of amphibian cathelicidin and its ability to influence immune cell function. This will expedite the potential utilization of amphibian antimicrobial peptides as therapeutic agents.

[The role of apolipoprotein C3 in the regulation of nonalcoholic fatty liver disease, glucose and lipid metabolism, and islet ß cell function].

As a member of the apolipoprotein C (ApoC) family with a relatively high content, ApoC3 plays a major role in the regulation of triglyceride metabolism, and plays an important role in the occurrence and development of cardiovascular diseases, glucose and lipid metabolism disorders. Nonalcoholic fatty liver disease (NAFLD) refers to the accumulation of a large amount of fat in the liver in the absence of a history of chronic alcohol consumption or other damage to the liver. A large number of previous studies have shown that there is a correlation between the gene polymorphism and high expression of ApoC3 and NAFLD. In the context of hypertriglyceridemia (HTG), this article reviews the relationship between ApoC3 and NAFLD, glucose and lipid metabolism, and islet ß cell function, showing that ApoC3 can not only inhibit lipoprotein lipase (LPL) and hepatic lipase (HL) activity, delay the decomposition of triglyceride in plasma to maintain the body's energy metabolism during fasting, but also be significantly increased under insulin resistance, prompting the liver to secrete a large amount of very low-density lipoprotein (VLDL) to induce HTG. Therefore, targeting and inhibiting ApoC3 might become a new approach to treat HTG. Increasing evidence suggests that ApoC3 does not appear to be an independent "contributor" to NAFLD. Similarly, our previous studies have shown that ApoC3 is not an independent factor triggering islet ß cell dysfunction in ApoC3 transgenic mice, but in a state of excess nutrition, HTG triggered by ApoC3 high expression may exacerbate the effects of hyperglycemia and insulin resistance on islet ß cell function, and the underlying mechanism remains to be further discussed.

Mefunidone ameliorates lipopolysaccharide-induced acute lung injury through inhibiting MAPK signaling pathway and enhancing Nrf2 pathway.

BACKGROUND AND OBJECTIVE: Acute lung injury (ALI) is a life-threatening disease which has high mortality and lacks effective pharmacological treatments. Excessive inflammation and oxidative stress are the key pathogenesis of ALI. Mefunidone (MFD), a novel small molecule compound, displayed anti-inflammation and anti-oxidative stress effects on streptozocin (STZ) and db/db mice in our previous studies. In this study, we aimed to investigate the effects of MFD on lipopolysaccharide (LPS)-induced ALI and explore the potential molecular mechanisms. METHODS: We investigated the effects of MFD on LPS-induced ALI mouse model and LPS-stimulated immortalized mouse bone marrow-derived macrophages (iBMDMs). RESULTS: MFD could alleviate pulmonary structure disorder and attenuate pulmonary neutrophils infiltration induced by LPS. MFD could also decreased proinflammatory cytokines release and reduce reactive oxygen species (ROS) generation stimulated by LPS. Further, MFD could significantly reduce LPS-induced phosphorylation levels of mitogen-activated protein kinase (MAPK), increase expression of nuclear factor-erythroid 2 related factor 2 (Nrf2) and restore the expressions of antioxidant enzymes. CONCLUSION: Our results firstly supported that MFD effectively protected LPS-induced ALI against inflammation and oxidative stress through inhibiting MAPK signaling pathway and activating Nrf2 pathway.

Repeated inhalation of sevoflurane inhibits the information transmission of Purkinje cells and delays motor development via the GABAA receptor ε subunit in neonatal mice.

General anesthesia is widely used in pediatric surgery, although the influence of general anesthesia on cerebellar information transmission and motor function is unclear. In the present study, neonatal mice received repeated inhalation of sevoflurane, and electrophysiological alterations in Purkinje cells (PCs) and the development of motor functions were detected. In addition, γ­aminobutyric acidA receptor ε (GABAA­R ε) subunit knockout mice were used to investigate the mechanism of action of sevoflurane on cerebellar function. In the neonatal mice, the field potential response of PCs induced by sensory stimulation and the motor function indices were markedly inhibited by sevoflurane, and the inhibitory effect was positively associated with the number of repetitions of anesthesia. In additional the GABAA­R ε subunit level of PCs was promoted by sevoflurane in a dose­dependent manner, and the inhibitory effects of sevoflurane on PC field potential response and motor function were alleviated in GABAA­R ε subunit knockout mice. The GABAA­R ε subunit was activated by sevoflurane, leading to inhibition of sensory information transmission in the cerebellar cortex, field potential responses of PCs and the development of cerebellar motor function. The present study provided experimental evidence for the safe usage of sevoflurane in clinical anesthesia, and suggested that GABAA­R ε subunit antagonists may be considered for combined application with general anesthesia with repeated inhalation of sevoflurane, for adverse effect prevention in the clinic.

Elevated urinary levels of cystatin C and neutrophil gelatinase-associated lipocalin in Henoch-Schönlein purpura patients with renal involvement.

Henoch-Schönlein purpura (HSP) is a commonest systemic vasculitis in childhood. The long-term prognosis of HSP is determined by the degree of renal involvement. The aim of this study is to search novel clinically applicable biomarkers to evaluate renal involvement in HSP patients. 20 bio-indexes in urine samples were simultaneously screened by antibody array assay. We indicated that urinary levels of cystatin C (Cys C) and neutrophil gelatinase-associated lipocalin (NGAL) in HSP patients with renal involvement were significantly higher than those without renal involvement and healthy controls. Furthermore, ELISA was used to analyze urinary Cys C and NGAL levels in HSP patients with or without renal involvement, atopic dermatitis (AD) patients and healthy controls. Our results demonstrated that urinary Cys C and NGAL levels in HSP patients with renal involvement were significantly elevated, when compared with those without renal involvement, AD patients and control subjects. In addition, by receiver operating characteristic (ROC) curve analysis, we demonstrated that the area under the ROC curve of NGAL (0.789) was larger than that of Cys C (0.692). Taken together, we show firstly that urinary Cys C and NGAL levels is abnormally elevated in HSP patients with renal involvement. We suggest that urinary Cys C and NGAL are novel useful biomarkers of renal involvement in HSP patients.

Differential patterns of insulin secretion and sensitivity in patients with type 2 diabetes mellitus and nonalcoholic fatty liver disease versus patients with type 2 diabetes mellitus alone.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) and type 2 diabetes mellitus (T2DM) often coexist and have adverse outcomes. The aim of our study was to elucidate metabolic abnormalities in patients with DM-NAFLD versus those with T2DM alone. METHODS: Patients were divided into two groups: 26 T2DM patients with NAFLD and 26 gender-, age-, and body mass index-matched patients with T2DM alone. Patients took a 75-g oral glucose tolerance test (OGTT), which measured serum insulin and C-peptide (C-p) levels at baseline (0 min), 30 min, 60 min, and 120 min after glucose challenge. RESULTS: Patients with DM-NAFLD or T2DM alone had similar blood glucose levels. ß-cell hypersecretion was more obvious in patients with DM-NAFLD. In addition, fasting, early-phase, and late-phase C-peptide levels were significantly increased in patients with DM-NAFLD (ΔC-p 0-30 min, P < 0.05; Area Under the Curve (AUC) C-p/PG 30-120 min ratio, P < 0.01; and AUC C-p 30-120 min, P < 0.01). Hepatic and extrahepatic insulin resistance during the OGTT did not differ significantly between groups. Hepatic insulin sensitivity independently contributed to the early phase (0-30 min) of the OGTT in patients with T2DM and NAFLD, whereas a significant deficit in late insulin secretion independently contributed to the 30-120 min glucose status in patients with T2DM only. CONCLUSIONS: In patients with similar levels of insulin resistance and hyperglycemia, DM-NAFLD was associated with higher serum insulin levels than T2DM alone. Hyperinsulinemia is caused mainly by ß-cell hypersecretion. The present study demonstrates pathophysiological differences in mechanisms of insulin resistance in patients with DM-NAFLD versus T2DM alone.

Low bone mineral density in chinese adults with nonalcoholic Fatty liver disease.

Aim. To investigate bone metabolic characteristics in Chinese adults with nonalcoholic fatty liver disease (NAFLD). Methods. A total of 224 patients (99 males and 125 postmenopausal females) were recruited and divided into 4 groups: males without NAFLD, males with NAFLD, females without NAFLD, and females with NAFLD. Bone mineral density (BMD) was evaluated according to body mass index (BMI), waist circumference (WC), and serum biomarkers. ß cell function was evaluated by HOMA2%B, HOMA2%S, and HOMA2IR. Results. Males in the NAFLD group had lower BMD of the right hip and the femoral neck (0.852 ± 0.117 versus 0.930 ± 0.123, P = 0.002; 0.736 ± 0.119 versus 0.812 ± 0.132, P = 0.004), and females had lower BMD of the right hip (0.725 ± 0.141 versus 0.805 ± 0.145, P = 0.002) even after adjusted for weight, BMI, waist, HDL, and ALT. There was no significant difference in bone metabolic markers between patients with and without NAFLD. NAFLD was an important factor that affected the bone; moreover, the effect attenuated when HOMA2IR entered into the model (R (2) = 0.160, ß = -0.172, and P = 0.008). Conclusions. NAFLD exerts a detrimental effect on BMD in both males and females. Insulin resistance may play an important role in this pathophysiological process.

A novel semisynthetic molecule icaritin stimulates osteogenic differentiation and inhibits adipogenesis of mesenchymal stem cells.

BACKGROUND: We previously reported that the constitutional flavonoid glycosides derived from herb Epimedium (EF, composed of seven flavonoid compounds with common nuclear stem) exerted beneficial effects on the bone, including promoting bone formation and inhibiting bone marrow fat deposition. Recent in vivo study showed that Icaritin was a common metabolite of these constitutional flavonoid glycosides, indicating that Icaritin is a bioactive compound. The present study was designed to investigate whether Icaritin could promote osteogenic differentiation and suppress adipogenic differentiation of marrow mesenchymal stem cells (MSCs). METHODS: Primary MSCs were harvested from adult mice and exposed to Icaritin to evaluate whether it could promote osteogenesis and suppress adipogenesis using the following assays: determination of alkaline phosphatase (ALP) activity and mineralization; mRNA expression of osteogenic differentiation marker Runx2; osteocalcin and bone sialoprotein (BSP) by RT-PCR; quantification of adipocyte-like cells by Oil Red O staining assay and mRNA expression for adipogenic differentiation markers peroxisome proliferator-activated receptor gamma (PPARγ); adipocyte fatty acid binding protein (aP2) and lipoprotein lipase (LPL) by RT-PCR. For the underlying mechanism, glycogen synthase kinase-3beta (GSK3ß) and ß-catenin were also explored by western blotting. RESULTS: Icaritin promoted osteogenic differentiation and maturation of MSCs as indicated by increased mRNA expression for Runx2, osteocalcin and BSP, and enhanced ALP activity and mineralization; Icaritin inhibited adipogenic differentiation, as indicated by decreased mRNA expression for PPARγ, LPL, aP2, and suppressed formation of adipocyte-like cells; Icaritin inactivated GSK3ß and suppressed PPARγ expression when promoting osteogenesis and suppressing adipogenesis of MSCs. CONCLUSION: This was the first study demonstrating that the novel semisynthetic molecule Icaritin could stimulate osteogenic differentiation and inhibit adipogenesis of MSCs, which was associated with the suppression of GSK3ß and PPARγ.

Pathomorphological changes of bone marrow adipocytes in process of steroid-associated osteonecrosis.

OBJECTIVE: The role of extravascular fat deposition in pathogenesis of steroid-associated osteonecrosis (ON) still remains unclear. This study aimed to explore the pathomorphological changes of bone marrow adipocytes over time in a rabbit ON model. METHODS: Thirty-two adult rabbits were divided into control group (n=16) and steroid group (n=16). Rabbits in the steroid group were injected with venous lipopolysaccharide once and intramuscular methylprednisolone trice to induce ON. Rabbits in the control group were treated with normal saline of equal volume. 2 weeks (early stage; n=8) and 4 weeks (late stage; n=8) after the last steroid injection, animals were sacrificed, and bilateral femora were harvested. The density, diameter and area of bone marrow adipocytes were determined by histomorphometry, and ON was evaluated histopathologically. RESULTS: The adipocyte density in steroid group increased by 67.1% and 54.4% at week 2 and week 4, respectively, when compared with control group, but there was no significant difference between week 2 and week 4. The adipocyte diameter in the steroid group at week 4 was significantly larger than that in the control group, but the adipocyte diameter in the steroid group at week 2 was slightly smaller than that in the control group. The adipocyte area in the steroid group increased by 44% and 83.4% at week 2 and week 4, respectively, when compared with the control group, and the adipocyte area in the steroid group at week 4 was markedly larger than that at week 2. In the control group, there were a largest number of adipocytes with 40-50 µm in diameter. When compared with the control group, most of increased adipocytes in the steroid group at week 2 were 30-40 µm in diameter, and those at week 4 were 50-60 µm in diameter. In the steroid group, histopathological examination showed ON was found in 25% (2/8) of rabbits at week 2 and 87.5% (7/8) of rabbits at week 4. CONCLUSION: In the process of ON, extravascular fat deposition is characterized by increased small adipocytes at the early stage and hypertrophy of adipocytes at the late stage.

The skeletal effects of thiazolidinedione and metformin on insulin-resistant mice.

To explore the skeletal effects and the potential underlying mechanisms of treatment with two thiazolidinediones (rosiglitazone and pioglitazone) or metformin in insulin-resistant mice, 24 female, 12-week-old C57BL6J ob/ob mice were evaluated according to the following treatment groups for 6 weeks: placebo group, pioglitazone group (Pio), rosiglitazone group (Rosi), and metformin group (Met). Bone mineral density (BMD), bone microarchitecture, bone histomorphometry, and expression of three phenotype-specific gene markers, including bone morphogenetic protein 2 (Bmp2), runt-related transcription factor 2 (Runx2), and fatty acid-binding protein 4 (Fabp4), were compared across the four groups. At the femur, the Met group had the highest BMD (0.084 ± 0.001 g/cm(2)) and trabecular bone volume/total volume (0.644 ± 0.018 %) and the lowest trabecular spacing (Tb.Sp.) (0.143 ± 0.008 µm), whereas the Rosi group had lower BMD (0.076 ± 0.003 g/cm(2)) and a relatively higher degree of Tb.Sp. (0.173 ± 0.024 µm). A histomorphometric analysis revealed that in the Rosi group the number of adipocytes was fourfold higher than in the placebo group and fivefold higher than in the Met group, whereas the highest osteoid width and mineral apposition rate were found in the Met group (49.88 ± 48.53 µm and 4.46 ± 1.72 µm/day). Furthermore, the Rosi group displayed the highest level of Fabp4 gene expression, which was accompanied by normal expression levels of Bmp2 and Runx2. Seemingly, metformin is a bone-friendly antidiabetic drug. Rosiglitazone had adverse effects on the skeleton at the trabecular bone even in insulin-resistant mice, whereas no evidence of adverse effects was found for pioglitazone.