Sorbitol Destroyed Intestinal Microfold Cells (M Cells) Development through Inhibition of PDE4-Mediated RANKL Expression.

Objective: Microfold cells (M cells) are specific intestinal epithelial cells for monitoring and transcytosis of antigens, microorganisms, and pathogens in the intestine. However, the mechanism for M-cell development remained elusive. Materials and Methods: Real-time polymerase chain reaction, immunofluorescence, and western blotting were performed to analyze the effect of sorbitol-regulated M-cell differentiation in vivo and in vitro, and luciferase and chromatin Immunoprecipitation were used to reveal the mechanism through which sorbitol-modulated M-cell differentiation. Results: Herein, in comparison to the mannitol group (control group), we found that intestinal M-cell development was inhibited in response to sorbitol treatment as evidenced by impaired enteroids accompanying with decreased early differentiation marker Annexin 5, Marcksl1, Spib, sox8, and mature M-cell marker glycoprotein 2 expression, which was attributed to downregulation of receptor activator of nuclear factor kappa-В ligand (RANKL) expression in vivo and in vitro. Mechanically, in the M-cell model, sorbitol stimulation caused a significant upregulation of phosphodiesterase 4 (PDE4) phosphorylation, leading to decreased protein kinase A (PKA)/cAMP-response element binding protein (CREB) activation, which further resulted in CREB retention in cytosolic and attenuated CREB binds to RANKL promoter to inhibit RANKL expression. Interestingly, endogenous PKA interacted with CREB, and this interaction was destroyed by sorbitol stimulation. Most importantly, inhibition of PDE4 by dipyridamole could rescue the inhibitory effect of sorbitol on intestinal enteroids and M-cell differentiation and mature in vivo and in vitro. Conclusion: These findings suggested that sorbitol suppressed intestinal enteroids and M-cell differentiation and matured through PDE4-mediated RANKL expression; targeting to inhibit PDE4 was sufficient to induce M-cell development.

In silico study of androgen receptor N-terminal domain and exploration of its modulators.

The androgen receptor (AR, Uniprot: P10275) signaling plays a key role in the progression of prostate cancer, various AR-related ligands have been reported to treat prostate cancer. However, some resistance mechanisms limited the treating effect of these ligands. Since DBD binding or the allosteric binding sites in LBD of AR may allow the circumvention of some drug resistance mechanisms, anti-resistance is expected especially through the NTD (N-terminal domain) targeting. What's more, studies have shown that compounds including EPI-001 and its derivatives which bind to the Tau-5 region on NTD could be promising molecules for AR-based therapeutics. Herein, we employed aMD (accelerated molecular dynamics) simulation to fold Tau-5 unit proteins into native structure correctly. Subsequently, based on the predicted structural features of Tau-5, the virtual screening was conducted to discover new compounds targeting AR-NTD. We picked up 8 compounds (according to their docking scores and partly similar structural consists as known AR ligands) and analyzed their interaction with Tau-5, compared with the positive control EPI-001, four of the pick-up compounds showed better glide scores. Interestingly, although compound 8 had a lower docking score, it consisted of a similar component as the ligand EIQPN and the amide derivatives, this predicts that compound 8 has also the potential to be modified into an excellent AR-NTD binding molecule. These 8 compounds were all commercially available and could be tested to check whether there was a hit compound to bind the AR-NTD and to regulate its bio-activities. Together, this study described an in silico VLS approach to discover AR-NTD ligands and provided more choices for developing AR-targeted therapies.Communicated by Ramaswamy H. Sarma.

Interferon-Gamma-Inducible Protein 16 Inhibits Hepatocellular Carcinoma via Interferon Regulatory Factor 3 on Chemosensitivity.

BACKGROUND AND AIM: Previous reports have suggested IFI16 as a tumor suppressor in hepatocellular carcinoma (HC). Nonetheless, the biological significance of IFI16 and its mechanism concerning resistance to cisplatin (DDP) in HC requires further exploration. METHODS: Samples of tumor and corresponding para-carcinoma tissues were acquired from patients with HC. Furthermore, DDP-resistant cell lines of HC, specifically HCC, Huh7 and Hepatoblastoma, HepG3, were generated by gradually increasing the concentration of DDP. Cell apoptosis and DNA damage were evaluated by utilizing flow cytometry assay and TUNEL staining. The interaction between IFI16 and interferon regulatory factor 3 (IRF3) proteins were analyzed using Co-Immunoprecipitation (Co-IP) assay. In vivo assays were conducted by establishing HC subcutaneous xenograft tumor models. RESULTS: The study found a reduction in IFI16 expression in both HC tissues and DDP-resistant HC cell lines. The binding of IFI16 to IRF3 regulated DNA damage-associated markers in vitro. Overexpression of IFI16 heightened the susceptibility of DDP-induced apoptosis and DNA damage, which was counteracted by IRF3 knockdown, while strengthened by IRF3 overexpression. Moreover, overexpression of IFI16 diminished in vivo DDP-resistant HC tumorigenicity. CONCLUSION: In summary, our findings suggest that IFI16 serves as a tumor suppressor in HC by promoting DNA damage via its interaction with IRF3, thereby reversing DDP resistance.

The novel mechanism of human norovirus induced diarrhea: Activation of PKD2 caused by HuNoVs destroyed AQP3 expression through AP2γ in intestinal epithelial cells.

Our previous work has demonstrated protein kinase D2 (PKD2) played a critical influence in experimental colitis in animal. However, the role of PKD2 in human norovirus (HuNoVs)-induced diarrhea remained unknown. Aquaporin 3 (AQP3) expression, a critical protein mediating diarrhea, was assessed by western blot, qRT-PCR in intestinal epithelial cells (IECs). Luciferase, IF, IP and ChIP assay were used to explore the mechanism through which HuNoVs regulated AQP3. Herein, we found that AQP3 expression was drastically decreased in IECs in response to VP1 transfection, the major capsid protein of HuNoVs, or HuNoVs infection. Mechanistically, HuNoVs triggered phosphorylation of PKD2 through TLR2/MyD88/IRAK4, which further inhibited AP2γ activation and nuclear translocation, leading to suppress AQP3 transactivation in IECs. Most importantly, PKD2 interacted with MyD88/IRAK4, and VP1 overexpression enhanced this complex form, which, in turn, to increase PKD2 phosphorylation. In addition, endogenous PKD2 interacted with AP2γ, and this interaction was enhanced in response to HuNoVs treatment, and subsequently resulting in AP2γ phosphorylation inhibition. Moreover, inhibition of PKD2 activation could reverse the inhibitory effect of HuNoVs on AQP3 expression. In summary, we established a novel mechanism that HuNoV inhibited AQP3 expression through TLR2/MyD88/IRAK4/PKD2 signaling pathway, targeting PKD2 activity could be a promising strategy for prevention of HuNoVs-induced gastroenteritis.

Vitronectin Destroyed Intestinal Epithelial Cell Differentiation through Activation of PDE4-Mediated Ferroptosis in Inflammatory Bowel Disease.

Objective: Vitronectin (VTN) has been reported to trigger cell pyroptosis to aggravate inflammation in our previous study. However, the function of VTN in inflammatory bowel disease (IBD) remains to be addressed. Methods: Real-time PCR and western blotting were performed to analyze VTN-regulated intestinal epithelial cell (IEC) differentiation through ferroptosis, and immunofluorescence (IF), luciferase, and chromatin immunoprecipitation were used to identify whether VTN-modulated ferroptosis is dependent on phosphodiesterase 4 (PDE4)/protein kinase A (PKA)/cyclic adenosine monophosphate-response element-binding protein (CREB) cascade pathway. In vivo experiment in mice and a pilot study in patients with IBD were used to confirm inhibition of PDE4-alleviated IECs ferroptosis, leading to cell differentiation during mucosal healing. Results: Herein, we found that caudal-related homeobox transcription factor 2-mediated IECs differentiation was impaired in response to VTN, which was attributed to enhanced ferroptosis characterized by decreased glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 expression. Inhibition of ferroptosis in IECs rescued the inhibitory effect of VTN on cell differentiation. Further analysis showed that VTN triggered phosphorylation of PDE4, leading to inhibit PKA/CREB activation and CREB nuclear translocation, which further reduced GPX4 transactivation. Endogenous PKA interacted with CREB, and this interaction was destroyed in response to VTN stimulation. What is more, overexpression of CREB in CaCO2 cells overcame the promotion of VTN on ferroptosis. Most importantly, inhibition of PDE4 by roflumilast or dipyridamole could alleviate dextran sulfate sodium-induced colitis in mice and in a pilot clinical study confirmed by IF. Conclusions: These findings demonstrated that highly expressed VTN disrupted IECs differentiation through PDE4-mediated ferroptosis in IBD, suggesting targeting PDE4 could be a promising therapeutic strategy for patients with IBD.

Risk factors for hepatic hydrothorax in patients with cirrhosis: a clinical retrospective study.

Aims and background: Hepatic hydrothorax, which presents as an unexplained pleural effusion, is one of the important complications in patients with end-stage cirrhosis. It has a significant correlation with prognosis and mortality. The aim of this clinical study was to detect the risk factors for hepatic hydrothorax in patients with cirrhosis and to better understand potentially life-threatening complications. Methods: Retrospectively, 978 cirrhotic patients who were hospitalized at the Shandong Public Health Clinical Center from 2013 to 2021 were involved in this study. They were divided into the observation group and the control group based on the presence of hepatic hydrothorax. The epidemiological, clinical, laboratory, and radiological characteristics of the patients were collected and analyzed. ROC curves were used to evaluate the forecasting ability of the candidate model. Furthermore, 487 cases in the experimental group were divided into left, right, and bilateral groups, and the data were analyzed. Results: The patients in the observation group had a higher proportion of upper gastrointestinal bleeding (UGIB), a history of spleen surgery, and a higher model for end-stage liver disease (MELD) scores compared with the control group. The width of the portal vein (PVW) (P = 0.022), prothrombin activity (PTA) (P = 0.012), D-dimer (P = 0.010), immunoglobulin G (IgG) (P = 0.007), high-density lipoprotein cholesterol (HDL) (P = 0.022), and the MELD score were significantly associated with the occurrence of the hepatic hydrothorax. The AUC of the candidate model was 0.805 (P < 0.001, 95% CI = 0.758-0.851). Portal vein thrombosis was more common in bilateral pleural effusion compared with the left and right sides (P = 0.018). Conclusion: The occurrence of hepatic hydrothorax has a close relationship with lower HDL, PTA, and higher PVW, D-dimer, IgG, and MELD scores. Portal vein thrombosis is more common in cirrhotic patients with bilateral pleural effusion compared to those with unilateral pleural effusion.

Helicobacter pylori promotes gastric intestinal metaplasia through activation of IRF3-mediated kynurenine pathway.

BACKGROUND: Metabolic reprogramming is a critical event for cell fate and function, making it an attractive target for clinical therapy. The function of metabolic reprogramming in Helicobacter pylori (H. pylori)-infected gastric intestinal metaplasia remained to be identified. METHODS: Xanthurenic acid (XA) was measured in gastric cancer cells treated with H. pylori or H. pylori virulence factor, respectively, and qPCR and WB were performed to detect CDX2 and key metabolic enzymes expression. A subcellular fractionation approach, luciferase and ChIP combined with immunofluorescence were applied to reveal the mechanism underlying H. pylori mediated kynurenine pathway in intestinal metaplasia in vivo and in vitro. RESULTS: Herein, we, for the first time, demonstrated that H. pylori contributed to gastric intestinal metaplasia characterized by enhanced Caudal-related homeobox transcription factor-2 (CDX2) and mucin2 (MUC2) expression, which was attributed to activation of kynurenine pathway. H. pylori promoted kynurenine aminotransferase II (KAT2)-mediated kynurenine pathway of tryptophan metabolism, leading to XA production, which further induced CDX2 expression in gastric epithelial cells. Mechanically, H. pylori activated cyclic guanylate adenylate synthase (cGAS)-interferon regulatory factor 3 (IRF3) pathway in gastric epithelial cells, leading to enhance IRF3 nuclear translocation and the binding of IRF3 to KAT2 promoter. Inhibition of KAT2 could significantly reverse the effect of H. pylori on CDX2 expression. Also, the rescue phenomenon was observed in gastric epithelial cells treated with H. pylori after IRF3 inhibition in vitro and in vivo. Most importantly, phospho-IRF3 was confirmed to be a clinical positive relationship with CDX2. CONCLUSION: These finding suggested H. pylori contributed to gastric intestinal metaplasia through KAT2-mediated kynurenine pathway of tryptophan metabolism via cGAS-IRF3 signaling, targeting the kynurenine pathway could be a promising strategy to prevent gastric intestinal metaplasia caused by H. pylori infection. Video Abstract.

Injectable self-assembled dual-crosslinked alginate/recombinant collagen-based hydrogel for endometrium regeneration.

The disadvantages of mainstream therapies for endometrial injury are difficult to resolve, herein, we suggest an omnibearing improvement strategy by introducing an injectable multifunctional self-assembled dual-crosslinked sodium alginate/recombinant collagen hydrogel. The hydrogel possessed a reversible and dynamic double network based on dynamic covalent bonds and ionic interactions, which also contributed to excellent capability in viscosity and injectability. Moreover, it was also biodegradable with a suitable speed, giving off active ingredients during the degradation process and eventually disappearing completely. In vitro tests exhibited that the hydrogel was biocompatible and able to enhance endometrial stromal cells viability. These features synergistically promoted cell multiplication and maintenance of endometrial hormone homeostasis, which accelerated endometrial matrix regeneration and structural reconstruction after severe injury in vivo. Furthermore, we explored the interrelation between the hydrogel characteristics, endometrial structure, and postoperative uterine recovery, which would benefit deep research on regulation of uterine repair mechanism and optimization of hydrogel materials. The injectable hydrogel could achieve favourable therapeutic efficacy without the need of exogenous hormones or cells, which would be of clinical value in endometrium regeneration.

Nanovaccine that activates the NLRP3 inflammasome enhances tumor specific activation of anti-cancer immunity.

Neoantigen cancer vaccines that target tumor specific mutations are emerging as a promising modality for cancer immunotherapy. To date, various approaches have been adopted to enhance efficacy of these therapies, but the low immunogenicity of neoantigens has hindered clinical application. To address this challenge, we developed a polymeric nanovaccine platform that activates the NLRP3 inflammasome, a key immunological signaling pathway in pathogen recognition and clearance. The nanovaccine is comprised of a poly (orthoester) scaffold engrafted with a small-molecule TLR7/8 agonist and an endosomal escape peptide that facilitates lysosomal rupture and NLRP3 inflammasome activation. Upon solvent transfer, the polymer self-assembles with neoantigens to form ∼50 nm nanoparticles that facilitate co-delivery to antigen-presenting cells. This polymeric activator of the inflammasome (PAI) was found to induce potent antigen-specific CD8+ T cell responses characterized by IFN-γ and GranzymeB secretion. Moreover, in combination with immune checkpoint blockade therapy, the nanovaccine stimulated robust anti-tumor immune responses against established tumors in EG.7-OVA, B16·F10, and CT-26 models. Results from our studies indicate that NLRP3 inflammasome activating nanovaccines demonstrate promise for development as a robust platform to enhance immunogenicity of neoantigen therapies.

An overview on the exploring the interaction of inorganic nanoparticles with microtubules for the advancement of cancer therapeutics.

Targeting microtubules (MTs), dynamic and stable proteins in cells, by different ligands have been reported to be a potential strategy to combat cancer cells. Inorganic nanoparticles (NPs) have been widely used as anticancer, antibacterial and free radical scavenging agents, where they come in contact with biological macromolecules. The interaction between the NPs and biological macromolecules like MTs frequently occurs through different mechanisms. A prerequisite for a detailed exploration of MT structures and functions for biomedical applications like cancer therapy is to investigate profoundly the mechanisms involved in MT-NP interactions, for which the full explanation and characterization of the parameters that are responsible for the formation of a NP-protein complex are crucial. Therefore, in view of the fact that the goal of the rational NP-based future drug design and new therapies is to rely on the information of the structural details and protein-NPs binding mechanisms to manipulate the process of developing new potential drugs, a comprehensive investigation of the essence of the molecular recognition/interaction is also of considerable importance. In the present review, first, the microtubule (MT) structure and its binding sites upon interaction with MT stabilizing agents (MSAs) and MT destabilizing agents (MDAs) are introduced and rationalized. Next, MT targeting in cancer therapy and interaction of NPs with MTs are discussed. Furthermore, interaction of NPs with proteins and the manipulation of protein corona (PC), experimental techniques and direct interaction of NPs with MTs, are discussed, and finally the challenges and future perspective of the field are introduced. We envision this review can provide useful information on the manipulation of the MT lattice for the progress of cancer nanomedicine.

Pathogen-like Nanoassemblies of Covalently Linked TLR Agonists Enhance CD8 and NK Cell-Mediated Antitumor Immunity.

Therapies based on Toll Like Receptor agonists (TLRa) are emerging as a promising modality for cancer immunotherapy to recruit antitumor T-cells in unresponsive immunologically "cold" tumors. Often, combinations of agonists are employed to synergistically enhance efficacy. However, low efficacy and severe toxicities deter these TLR-based therapeutics from further clinical applications. Studies have suggested that the rapid systemic diffusion of agonists to nontarget tissues is the primary cause. To address this challenge, we developed supramolecular nanotherapeutics of covalently linked TLRas for multivalent, synergistic interactions by drawing inspiration from immune recognition of pathogens. This new nanotherapeutic increased stimulation of key pro-inflammatory cytokines and remarkably enhanced CD8 and NK cell-mediated antitumor response while exhibiting ultralow off-target toxicity in an aggressive B16.F10 tumor model. Results from our studies thereby indicate that such supramolecular immune-agonist therapeutics may be further developed as a viable treatment modality for cancer immunotherapy.

Low preoperative serum ALB level is independently associated with poor overall survival in endometrial cancer patients.

Aim: To reveal the prognostic significance of serum albumin (ALB) concentration in endometrial cancer (EC) patients in China. Patients & methods: 345 EC patients were enrolled in a single center, and the preoperative serum ALB concentration were measured. Kaplan-Meier curve analysis and Cox proportional hazards regression model were performed to evaluate the associations between ALB concentration and overall survival (OS) of EC patients. Results: The EC patients with lower preoperative serum ALB concentration exhibited a significantly poorer OS (p < 0.05). Univariate analysis and multivariate analysis indicated that serum ALB concentration was an independent prognostic factor of unfavorable OS for EC patients. Conclusion: Our results showing that ALB concentration may serve as an independent prognostic factor for EC patients.

Bioinformatics analysis of LINC00426 expression in lung cancer and its correlation with patients' prognosis.

BACKGROUND: To investigate the expression of long noncoding RNA (lncRNA) LINC00426 (long intergenic nonprotein coding RNA 426) in non-small cell lung cancer (NSCLC) patients and its correlation with their prognosis. METHODS: The expression of long noncoding RNA LINC00426 of non-small cell lung cancer (NSCLC) in The Cancer Genome Atlas (TCGA) database was screened. According to the expression level of LINC00426 in tumor tissue of NSCLC patients, the patients were divided into high and low LINC00426 expression groups. The correlation between LINC00426 expression group and the prognosis of the patient was analyzed by log-rank test. A total of 72 NSCLC patients who had undergone surgery were retrospectively included in this study. LINC00426 relative expression of tumor and normal lung tissue of the included 72 NSCLC patients were examined by real-time quantitative PCR assay. The correlation between LINC00426 expression and the patients' clinical characteristics were also evaluated. RESULTS: LINC00426 relative expression was not statistically different between cancer and normal tissue (P > 0.05) of NSCLC patients in the TCGA database. The amplification and deep deletion mutation of LINC00426 gene was found in 0.5% of NSCLC patients. The overall survival (OS) of the LINC00426 high expression group was significantly higher than that of the low expression group (HR = 0.81, P = 0.044), while there was no significant difference between the high and low expression group (HR = 0.97, P = 0.82) for disease-free survival (DFS). LINC0042646 expression level was elevated in 46 cases in normal lung tissue compared to the tumor tissue of the 72 NSCLC patients. LINC0042646 expression level was significantly correlated with the clinical stage (P < 0.05). CONCLUSION: Long noncoding RNA LINC00426 was downregulated in the tumor tissue of NSCLC patients and correlated with poor prognosis.

Discovery of aminopyridine-containing spiro derivatives as EGFR mutations inhibitors.

Neratinib is an oral pan HER inhibitor, that irreversibly inhibits EGFR and HER2 and was proven to be effective against multiple EGFR mutations. In previous study, we reported spiro [indoline-3, 4'-piperidine]-2-ones as anticancer agents. In this study, we designed aminopyridine-containing spiro [indoline-3,4'-piperidine] derivatives A1-A4 using Neratinib and spiro [indoline-3, 4'-piperidine]-2-one compound patented as lead structure, then replaced piperidine with cyclopropane to obtain B1-B7 and replaced indoline with benzmorpholine to get C1-C4 and D1-D2. We synthesized these compounds and evaluated their residual activities under 0.5 M drug concentration on EGFR and ERBB2. Most of compounds showed stronger inhibition on EGFR-wt and ERBB2, in which A1-A4 showed excellent inhibitory activity with inhibition percentage on EGFR-wt kinase of 7%, 6%, 19%, 27%, respectively and 9%, 5%, 12%, 34% on ERBB2 kinase compared with 2% and 6% of Neratinib.

Total phenolic, flavonoid content, and antioxidant activity of bulbs, leaves, and flowers made from Eleutherine bulbosa (Mill.) Urb.

In the current investigation, total phenols and flavonoids contents of Eleutherine bulbosa (Mill.) Urb. bulbs, leaves, and flowers were quantified by Folin-Ciocalteu's and borohydride/chloroquinone methods, respectively. Antioxidant activity of the plant extracts was evaluated by means of peroxide scavenging capacity assay and by cell antioxidation method. Antioxidant activity of E. bulbosa bulbs, leaves, and flowers was correlated with total phenols and flavonoids. The total phenols and flavonoids of the bulbs of E. bulbosa were higher than leaves and flower and its antioxidant activity was also stronger than leaves and flowers of E. bulbosa. The higher content of flavonoids or total phenols, the stronger the antioxidant capacity in vitro. The antioxidant activity of E. bulbosa extract showed it's certain nutritional value and therefore had the potential as a source of natural antioxidants.

IFI16 restoration in hepatocellular carcinoma induces tumour inhibition via activation of p53 signals and inflammasome.

OBJECTIVE: Interferon-inducible 16 (IFI16) is known to involve in p53-dependent tumour suppression and also the formation of inflammasome, which function, however, remains controversy during carcinogenesis as a pattern recognition receptor for tumour death-derived free DNA. In this study, we investigated the anti-tumour role of IFI16 in hepatocellular carcinoma (HCC). MATERIALS AND METHODS: Hepatocellular carcinoma tissues (n = 20) and corresponding non-neoplastic tissues (n = 20) were collected to determine the expression of IFI16. After the transfection of pcDNA3.1-IFI16 into Huh7 and SMMC7721 cells in vitro, the influence of IFI16 overexpression on cell vitality, colony formation, apoptosis and migration were analysed. The role effect of IFI16 in vivo was further investigated. RESULTS: The expression of IFI16 was significantly decreased in tumour tissues and cell lines. Overexpression of IFI16 induced decrease of cell vitality, colony formation and increased apoptosis with impaired ability of migration. Mechanistically, IFI16 could activate p53 at Ser15 to up-regulate the p21WAF1/CIP1 level to inhibit tumour growth and migration, which was restored by the p53 inhibitor Pifithrin-α (20 µmol/L). Moreover, IFI16-induced tumour cell death promoted the recruitment of inflammasome complex to enhance tumour inhibition, but the caspase-1 inhibitor Ac-YVAD-CMK (50 µmol/L) could suppress this process in HCC. The results in vivo indicated that restored expression of IFI16 in tumour cells effectively promote tumour regression, which could be partly abrogated by the inhibition of activation of p53 signals or induced inflammasome. CONCLUSION: IFI16 is a tumour suppressor in HCC via activation of p53 signals and inflammasome.

CGRP may regulate bone metabolism through stimulating osteoblast differentiation and inhibiting osteoclast formation.

Calcitonin-gene-related peptide (CGRP) is a neuropeptide, which is widely distributed throughout the central and peripheral nervous systems. Numerous mechanisms underlying the action of CGRP in osteoblast-associated cells have been suggested for bone growth and metabolism. The present study was designed to closely investigate the osteoblast­ and osteoclast-associated mechanisms of the effect of CGRP administration on bone metabolism in primary osteoblasts. Primary osteoblasts were obtained from newborn rabbit calvaria and incubated with different concentrations of human CGRP (hCGRP), hCGRP and hCGRP (8­37), or without treatment as a control. Intracellular calcium (Ca2+) and cyclic adenosine monophosphate (cAMP) were detected following treatment, as well as the expression levels of osteoblast differentiation markers, including activating transcription factor­4 (ATF4) and osteocalcin (OC), and receptor activator of nuclear factor κB ligand (RANKL) and osteoprotegerin (OPG). The isolated primary osteoblasts were found to stain positively for ALP. hCGRP treatment had no significant effect on transient intracellular Ca2+ in the osteoblasts. Treatment of the osteoblasts with hCGRP led to elevations in the expression levels of cAMP, ATF4 and OPG, and downregulation in the expression of RANKL, in a dose­dependent manner. These effects were markedly reversed by the addition of hCGRP (8­37). The results of the present study demonstrated that CGRP administration not only stimulated osteoblast differentiation, as demonstrated by upregulated expression levels of ATF4 and OC in the hCGRP­treated osteoblasts, but also inhibited OPG/RANKL­regulated osteoclastogenesis. CGRP may act as a modulator of bone metabolism through osteoblast and osteoclast-associated mechanisms, which result in osteoblast formation with subsequent activation of bone formation.

Quantitative assessment of the association between Fas/FasL gene polymorphism and susceptibility to esophageal carcinoma in a north Chinese population.

The case-control study aims to investigate the association of Fas and FasL genetic polymorphisms (Fas-670A/G (rs1800682), Fas-1377G/A (rs2234767) and FasL-844T/C (rs763110)) with esophageal carcinoma susceptibility in a north Chinese population. A total of 204 patients with esophageal carcinoma and 248 healthy controls were enrolled from Henan, China and genotyped by the polymerase chain reaction and restriction fragment length polymorphism method. There were no significant differences in distributions of their genotypes frequencies between patients and controls in Fas-670A/G, Fas-1377G/A and FasL-844T/C polymorphisms (P > 0.05). Stratified analysis showed that no significant association was found between esophageal carcinoma and gene polymorphisms of Fas-670 A/G, Fas-1377G/A, and FasL-844T/C (P > 0.05). Genetic polymorphisms in the death pathway genes Fas and FasL were not associated with risk of developing esophageal carcinoma in a north Chinese population.

Expression of AIM2 is correlated with increased inflammation in chronic hepatitis B patients.

BACKGROUND: The absent in melanoma 2 (AIM2), a cytosolic dsDNA inflammasome, can be activated by viral DNA to trigger caspase-1. Its role in immunopathology of chronic hepatitis B and C virus (HBV, HCV) infection is still largely unclear. In this study, the expression AIM2, and its downstream cytokines, caspase-1, IL-18 and IL-1ß, in liver tissue of patients with chronic hepatitis B and C (CHB, CHC) were investigated. METHODS: A total of 70 patients diagnosed with chronic hepatitis were enrolled, including 47 patients with CHB and 23 patients with CHC. A liver biopsy was taken from each patient, and immunohistochemistry was used to detect the expression of AIM2 and inflammatory factors caspase-1, IL-18, and IL-1ß in the biopsy specimens. The relationship between AIM2 expression and these inflammatory factors was analyzed. RESULTS: The expression of AIM2 in CHB patients (89.4 %) was significantly higher than in CHC patients (8.7 %), and among the CHB patients, the expression of AIM2 was significantly higher in the high HBV replication group (HBV DNA ≥ 1 × 10(5)copies/mL) than in the low HBV replication group (HBV DNA < 1 × 10(5)copies/mL). The expression of AIM2 was also correlated with HBV-associated inflammatory activity in CHB patients statistically. Additionally, AIM2 levels were positively correlated with the expression of caspase-1, IL-1ß and IL-18 in CHB patients, which implied that the AIM2 expression is directly correlated with the inflammatory activity associated with CHB. CONCLUSIONS: AIM2 upregulation may be a component of HBV immunopathology. The underlying mechanism and possible signal pathway warrant further study.

Conjugating an anticancer drug onto thiolated hyaluronic acid by acid liable hydrazone linkage for its gelation and dual stimuli-response release.

A prodrug gelation strategy was developed for the sustained and dual stimuli-response release of doxorubicin hydrochloride (DOX·HCl), a commonly used anticancer drug. For this purpose, the chemical conjugation of DOX·HCl onto thiolated hyaluronic acid (HA) was carried out by an acid liable hydrazone linkage and verified by (1)H NMR analyses. When exposed to the air, such a polysaccharide conjugate showed unique self-gelation ability in aqueous solution. The gelation time and extent depended mainly on the content of thiol groups on thiolated HA. The resultant hydrogel exhibited a dominant elastic response and a thixotropic property. In particular, it could release sustainably conjugated DOX·HCl in dual pH- and reduction-responsive modes. The cumulative drug release was found to be significantly accelerated under the conditions mimicking the intracellular environments of cancer cells. The in vitro cytotoxicity assays for the human nasopharyngeal carcinoma CNE2 cells treated with various release media confirmed the effectiveness of this conjugate hydrogel for cancer cell inhibition.