PHLDA1 contributes to hypoxic ischemic brain injury in neonatal rats via inhibiting FUNDC1-mediated mitophagy.

Hypoxia-ischemia (HI) is one of the main causes of neonatal brain injury. Mitophagy has been implicated in the degradation of damaged mitochondria and cell survival following neonatal brain HI injury. Pleckstrin homology-like domain family A member 1 (PHLDA1) plays vital roles in the progression of various disorders including the regulation of oxidative stress, the immune responses and apoptosis. In the present study we investigated the role of PHLDA1 in HI-induced neuronal injury and further explored the mechanisms underlying PHLDA1-regulated mitophagy in vivo and in vitro. HI model was established in newborn rats by ligation of the left common carotid artery plus exposure to an oxygen-deficient chamber with 8% O2 and 92% N2. In vitro studies were conducted in primary hippocampal neurons subjected to oxygen and glucose deprivation/-reoxygenation (OGD/R). We showed that the expression of PHLDA1 was significantly upregulated in the hippocampus of HI newborn rats and in OGD/R-treated primary neurons. Knockdown of PHLDA1 in neonatal rats via lentiviral vector not only significantly ameliorated HI-induced hippocampal neuronal injury but also markedly improved long-term cognitive function outcomes, whereas overexpression of PHLDA1 in neonatal rats via lentiviral vector aggravated these outcomes. PHLDA1 knockdown in primary neurons significantly reversed the reduction of cell viability and increase in intracellular reactive oxygen species (ROS) levels, and attenuated OGD-induced mitochondrial dysfunction, whereas overexpression of PHLDA1 decreased these parameters. In OGD/R-treated primary hippocampal neurons, we revealed that PHLDA1 knockdown enhanced mitophagy by activating FUNDC1, which was abolished by FUNDC1 knockdown or pretreatment with mitophagy inhibitor Mdivi-1 (25 µM). Notably, pretreatment with Mdivi-1 or the knockdown of FUNDC1 not only increased brain infarct volume, but also abolished the neuroprotective effect of PHLDA1 knockdown in HI newborn rats. Together, these results demonstrate that PHLDA1 contributes to neonatal HI-induced brain injury via inhibition of FUNDC1-mediated neuronal mitophagy.

Inhibition of USP10 induces myeloma cell apoptosis by promoting cyclin D3 degradation.

The cell cycle regulator cyclin D3 (CCND3) is highly expressed in multiple myeloma (MM) and it promotes MM cell proliferation. After a certain phase of cell cycle, CCND3 is rapidly degraded, which is essential for the strict control of MM cell cycle progress and proliferation. In the present study, we investigated the molecular mechanisms regulating CCND3 degradation in MM cells. By utilizing affinity purification-coupled tandem mass spectrometry, we identified the deubiquitinase USP10 interacting with CCND3 in human MM OPM2 and KMS11 cell lines. Furthermore, USP10 specifically prevented CCND3 from K48-linked polyubiquitination and proteasomal degradation, therefore enhancing its activity. We demonstrated that the N-terminal domain (aa. 1-205) of USP10 was dispensable for binding to and deubiquitinating CCND3. Although Thr283 was important for CCND3 activity, it was dispensable for CCND3 ubiquitination and stability modulated by USP10. By stabilizing CCND3, USP10 activated the CCND3/CDK4/6 signaling pathway, phosphorylated Rb, and upregulated CDK4, CDK6 and E2F-1 in OPM2 and KMS11 cells. Consistent with these findings, inhibition of USP10 by Spautin-1 resulted in accumulation of CCND3 with K48-linked polyubiquitination and degradation that synergized with Palbociclib, a CDK4/6 inhibitor, to induce MM cell apoptosis. In nude mice bearing myeloma xenografts with OPM2 and KMS11 cells, combined administration of Spautin-l and Palbociclib almost suppressed tumor growth within 30 days. This study thus identifies USP10 as the first deubiquitinase of CCND3 and also finds that targeting the USP10/CCND3/CDK4/6 axis may be a novel modality for the treatment of myeloma.

Totally laparoscopic total gastrectomy using the "enjoyable space" approach coupled with self-pulling and latter transection reconstruction versus laparoscopic-assisted total gastrectomy for upper gastric cancer: short-term outcomes.

Introduction: With the development of minimally invasive surgery in recent years, totally laparoscopic total gastrectomy (TLTG) has attracted more attention. Aim: To introduce the more comprehensive "enjoyable space" approach coupled with the self-pulling and latter transaction (SPLT) reconstruction technique to perform TLTG and investigate its safety and feasibility. Material and methods: Ninety-seven patients with primary upper gastric cancer underwent laparoscopic radical total gastrectomy between January 2020 and December 2020. Among these patients, 46 underwent laparoscopic-assisted total gastrectomy (LATG), and 51 underwent TLTG. We compared the clinicopathological characteristics, surgical outcomes and postoperative complications between the two groups. Results: There were no significant differences in the clinicopathological characteristics between the two groups (p > 0.05). However, the TLTG group had a slightly lower mean operative time and mean blood loss than the LATG group (p < 0.05 each). Although there were similarities in the mean times to first flatus, liquid diet, and soft diet, the duration of hospital stay was significantly reduced in the TLTG group (p < 0.05). No significant differences in overall complications and E-J-related complications were found between the two groups (15.2% vs. 25.4%, p > 0.05). Conclusions: TLTG is a safe and feasible procedure for treating upper gastric cancer. The enjoyable space approach in conjunction with SPLT reconstruction is an appropriate comprehensive technique with several advantages over LATG.

Prevalence and Clinical Predictors of Hyperuricemia in Chinese Bariatric Surgery Patients.

BACKGROUND: Despite a known significant association between hyperuricemia and obesity, this correlation in bariatric surgery patients remains unknown. OBJECTIVES: To evaluate the prevalence and predictors of pre- and postoperative hyperuricemia in Chinese bariatric surgery patients. METHODS: A retrospective study was performed in 333 bariatric surgery patients from our hospital. The clinical data was collected before surgery and at 3, 6, and 12 months postoperatively. Univariable and multivariate analyses were used for investigating the independent predictors of hyperuricemia and serum uric acid (SUA) change. RESULTS: Altogether, 62.9% of patients fulfilled the diagnostic criteria for hyperuricemia. The prevalence of hyperuricemia among males was 81.8% and 62.3% in the women. Multiple logistic regression analyses showed that age (OR = 0.951, 95%CI:0.926-0.976, P = 0.000), high-density lipoprotein cholesterol (HDL-c) (OR = 0.217, 95%CI:0.074-0.637, P = 0.005), γ-glutamyltransferase (γ-GT) (OR = 1.016, 95%CI:1.004-1.027, P = 0.006), and creatinine (Cr) (OR = 1.042, 95%CI: 1.017-1.067, P = 0.001) were independent predictors of hyperuricemia. SUA levels significantly declined in all patients from 443.1 ± 118.2 µmol/L before surgery to 370.1 + 113.4 µmol/L at 12 months after surgery. The prevalence of hyperuricemia also declined from 69.4% before surgery to 25.5% at 12 months. Multiple linear regression analyses confirmed that changes in Cr and body mass index (BMI) were independent predictors of a decrease in SUA levels, 12 months postoperatively. CONCLUSIONS: Hyperuricemia in Chinese bariatric surgery candidates are common, especially in males. Age, HDL-c, γ-GT and Cr were determined to be independent predictors of hyperuricemia. Bariatric surgery may effectively reduce the prevalence of hyperuricemia in this population, through postoperative weight loss and changes in creatinine following the procedure.

TRIM25 activates AKT/mTOR by inhibiting PTEN via K63-linked polyubiquitination in non-small cell lung cancer.

The PTEN/AKT/mTOR signaling pathway is frequently dysregulated in non-small cell lung cancer (NSCLC), but the mechanisms are not well-understood. The present study found that the ubiquitin ligase TRIM25 is highly expressed in NSCLC tissues and promotes NSCLC cell survival and tumor growth. Mechanistic studies revealed that TRIM25 binds to PTEN and mediates its K63-linked ubiquitination at K266. This modification prevents the plasma membrane translocation of PTEN and reduces its phosphatase activity therefore accumulating PI(3,4,5)P3. TRIM25 thus activates the AKT/mTOR signaling. Moreover, we found that the antibacterial nitroxoline can activate PTEN by reducing its K63-linked polyubiquitination and sensitizes NSCLC to cisplatin-induced apoptosis. This study thus identified a novel modulation on the PTEN signaling pathway by TRIM25 and provides a potential target for NSCLC treatment.

Atezolizumab alleviates the immunosuppression induced by PD­L1­positive neutrophils and improves the survival of mice during sepsis.

Atezolizumab can reduce immunosuppression caused by T lymphocyte apoptosis in various cancer types. The current study aimed to investigate whether this drug can also alleviate immunosuppression during sepsis. For that purpose, a C57BL/6 mouse sepsis model was generated. Mice were randomly assigned to three groups: Sham, cecal ligation and puncture (CLP) and atezolizumab groups. Atezolizumab was administered in vivo by intraperitoneal injection. The expression of programmed death ligand­1 (PD­L1) on neutrophils and programmed death­1 (PD­1) on T lymphocytes was evaluated, and endotoxin concentration, intestinal permeability, ileum histopathological score and tight junction protein expression were assessed to determine the extent of disease in each group. The rate of T lymphocyte apoptosis was determined to assess the effects of atezolizumab on T lymphocyte apoptosis in vivo and in vitro. Survival times were also recorded to compare mouse prognosis during sepsis. In the CLP group, the proportion of PD­L1+ neutrophils was significantly higher at 48, 72 and 96 h in blood, and at 24, 48, 72 and 96 h in bone marrow, compared with those of the sham group (P<0.05). The proportion of PD­1+ T lymphocytes was also upregulated at 72 h in blood. In the atezolizumab group, endotoxin concentration, intestinal permeability and ileum histopathological score were lower compared with those in the CLP group (P<0.05), whereas the expression of claudin­1 and occludin proteins on ileum was higher compared with that in the CLP group (P<0.05). Both in vivo and in vitro experiments indicated that the rate of T lymphocyte apoptosis following atezolizumab treatment was lower compared with that in the CLP group (P<0.05). Survival analysis demonstrated that mice in the atezolizumab group survived longer compared with those in the CLP group (P<0.05). The current study demonstrated that treatment with atezolizumab may be an effective method for treating immunosuppression induced by sepsis.

TP53-induced glycolysis and apoptosis regulator alleviates hypoxia/ischemia-induced microglial pyroptosis and ischemic brain damage.

Our previous studies have demonstrated that TP53-induced glycolysis and apoptosis regulator (TIGAR) can protect neurons after cerebral ischemia/reperfusion. However, the role of TIGAR in neonatal hypoxic-ischemic brain damage (HIBD) remains unknown. In the present study, 7-day-old Sprague-Dawley rat models of HIBD were established by permanent occlusion of the left common carotid artery followed by 2-hour hypoxia. At 6 days before induction of HIBD, a lentiviral vector containing short hairpin RNA of either TIGAR or gasdermin D (LV-sh_TIGAR or LV-sh_GSDMD) was injected into the left lateral ventricle and striatum. Highly aggressively proliferating immortalized (HAPI) microglial cell models of in vitro HIBD were established by 2-hour oxygen/glucose deprivation followed by 24-hour reoxygenation. Three days before in vitro HIBD induction, HAPI microglial cells were transfected with LV-sh_TIGAR or LV-sh_GSDMD. Our results showed that TIGAR expression was increased in the neonatal rat cortex after HIBD and in HAPI microglial cells after oxygen/glucose deprivation/reoxygenation. Lentivirus-mediated TIGAR knockdown in rats markedly worsened pyroptosis and brain damage after hypoxia/ischemia in vivo and in vitro. Application of exogenous nicotinamide adenine dinucleotide phosphate (NADPH) increased the NADPH level and the glutathione/oxidized glutathione ratio and decreased reactive oxygen species levels in HAPI microglial cells after oxygen/glucose deprivation/reoxygenation. Additionally, exogenous NADPH blocked the effects of TIGAR knockdown in neonatal HIBD in vivo and in vitro. These findings show that TIGAR can inhibit microglial pyroptosis and play a protective role in neonatal HIBD. The study was approved by the Animal Ethics Committee of Soochow University of China (approval No. 2017LW003) in 2017.

Laparoscopic D2 plus complete mesogastrium excision using the "enjoyable space" approach versus conventional D2 total gastrectomy for local advanced gastric cancer: short-term outcomes.

INTRODUCTION: Laparoscopy-assisted radical total gastrectomy is technically demanding. AIM: To introduce the "enjoyable space" approach to achieve D2 plus complete mesogastrium excision (CME) and to investigate its safety and feasibility. MATERIAL AND METHODS: Between January 2015 and December 2017, 165 patients with primary advanced upper gastric cancer underwent laparoscopy-assisted radical total gastrectomy. Among these patients, 81 underwent conventional D2 total gastrectomy (D2 group) and 84 underwent D2 plus CME total gastrectomy (D2 + CME group). Clinicopathological characteristics, surgical outcomes and postoperative complications were compared between the two groups. RESULTS: There were no significant differences between the two groups (p > 0.05) in clinicopathological characteristics. However, the D2 + CME group had a longer mean operative time, lower mean blood loss and slightly higher mean number of retrieved lymph nodes (LNs) than the D2 group (p < 0.05 each). The mean time to first flatus, liquid diet, and soft diet and the duration of hospital stay were similar between the two groups (p > 0.05 each). No significant difference in postoperative complication rates was found between the groups (16.0% vs. 9.5%, p > 0.05). CONCLUSIONS: The "enjoyable space" approach is an option to achieve D2 + CME, and its safety and feasibility over conventional method are confirmed with lower intraoperative blood loss and more harvested LNs.

Subacute Bowel Obstruction Secondary to an Obturator Hernia.

Obturator hernia is a rare abdominal wall hernia and is one of the rare causes of intestinal obstruction. Due to lack of specific clinical symptoms in the early stage, and the deep position of hernia absence of surface swelling, few cases are diagnosed definitively before surgery. Therefore, obturator hernia is still a serious disease in clinical practice, with high mortality and morbidity. Herein, we report an 85-year female who was hospitalised with bowel obstruction. Preoperative computed tomography scan was performed, which revealed an incarcerated obturator hernia preoperatively; and it was treated successfully. This report reviews the diagnosis and treatment of obturator hernia by describing its anatomy, clinical manifestations and clinical signs. Increased awareness and proper management of the disease will lead to a decline in mortality and morbidity.

RNF6 promotes myeloma cell proliferation and survival by inducing glucocorticoid receptor polyubiquitination.

RNF6, a RING-type ubiquitin ligase, has been identified as an oncogene in various cancers but its role in multiple myeloma (MM) remains elusive. In the present study we first showed that the expression levels of RNF6 in MM were significantly elevated compared with the bone marrow cells of healthy donors. Overexpression of RNF6 in LP1 and PRMI-8266 MM cell lines promoted cell proliferation, whereas knockdown of RNF6 led to apoptosis of MM cells. Furthermore, we revealed that RNF6, as a ubiquitin ligase, interacted with glucocorticoid receptor (GR) and induced its K63-linked polyubiquitination. Different from current knowledge, RNF6 increased GR stability at both endogenous and exogenous contexts. Such an action greatly promoted GR transcriptional activity, which was confirmed by luciferase assays and by the increased expression levels of prosurvival genes including Bcl-xL and Mcl-1, two typical downstream genes of the GR pathway. Consistent with these findings, ectopic expression of RNF6 in MM cells conferred resistance to dexamethasone, a typical anti-myeloma agent. In conclusion, we demonstrate that RNF6 promotes MM cell proliferation and survival by inducing atypical polyubiquitination to GR, and RNF6 could be a promising therapeutic target for the treatment of MM.

Expression and cellular function of long non－coding ＲNA SFTA1P in non－small cell lung cancer / 中华疾病控制杂志

@#Objective To investigate the expression of long non－coding ＲNA SFTA1P in non small cell lung cancer ( NSCLC) and its biological function in NSCLC cell lines． Methods Quantitative real time polymerase chain reaction( qＲT－PCＲ) was used to detect the expression of SFTA1P in 18 pairs of NSCLC tissues and adjacent normal tissues． The expression of SFTA1P was detected by qＲT－PCＲ in five different NSCLC cell lines ( A549，SPCA1，H460，H1975 and H1299) and one normal lung epithelial cell line ( HBE) ． The overexpression vector of SFTA1P was designed and constructed． The overex- pressed cell line was constructed by transfection，the effects of overexpression of SFTA1P on proliferation， invasion and migration of NSCLC cells were detected by CCK－8 assay and transwell assay． Ｒesults The expression of SFTA1P in NSCLC tissues was lower than that of adjacent normal tissues ( t = 2. 158，P = 0. 043) ． SFTA1P expression was detected in 5 strains of NSCLC cell lines and normal lung epithelial cell line． The expression of SFTA1P was the lowest in A549 and H460 cell lines ( t = 5. 769，P = 0. 004; t = 5. 772，P= 0. 004) ，and the highest in H1299 and H1975 cell lines ( t = 22. 248，P＜0. 001; t = 11. 814，P ＜0. 001) ． SFTA1P overexpression cell models were successfully constructed using A549 and H460 cell lines( all P＜0．05) ． The overexpression of SFTA1P could inhibit proliferation，invasion and migration of H460 and A549 cells ( ( all P ＜ 0． 05) ． Conclusions SFTA1P can affect the biological functions of NSCLC cells by inhibiting the proliferation，migration and invasion． SFTA1P may play a role as a tumor suppressor gene in tumorigenesis and development．

Retroperitoneoscopic distal pancreatectomy: a new surgical approach.

INTRODUCTION: The traditional laparoscopic surgery is difficult to deal with the deep lesions of the body and tail of the pancreas, which may damage the visceral organs of the abdominal cavity and cause abdominal adhesion and other related complications. AIM: This paper introduces the operation procedure of retroperitoneoscopy in pancreatic surgery, and evaluates its feasibility in clinical application. MATERIAL AND METHODS: Retrospective analysis was performed on patients with retroperitoneal pancreatectomy in our hospital. The anatomical features of the fascia, surgical plane composition and surgical pathway of the fascia of the retroperitoneoscopic pancreatectomy were observed during the operation, and the surgical safety and feasibility were analyzed. The following parameters were evaluated: operation time, blood loss, pancreatic fistula, postoperative gastro-intestinal recovery, hospital stay. RESULTS: All 3 patients had a smooth operation and no serious complications occurred. During retroperitoneal laparoscopic pancreatectomy, there is a vascularized plane between the posterior fascia of the pancreas and the prerenal fascia, which can avoid injury of the visceral organs and retroperitoneal vessels. The anterior renal fascia should be used as the posterior boundary of the safe separation plane. CONCLUSIONS: The surgical plane based on the anatomy of the fascia and interstitial dissection is the theoretical basis of modern surgery, which is safe, fast and effective. The inter-prerenal fascia plane is the correct and safe anatomical plane of posterior laparoscopic surgery.

Mebendazole elicits potent antimyeloma activity by inhibiting the USP5/c-Maf axis.

c-Maf is a critical oncogenic transcription factor that contributes to myelomagenesis. Our previous studies demonstrated that the deubiquitinase USP5 stabilizes c-Maf and promotes myeloma cell proliferation and survival; therefore, the USP5/c-Maf axis could be a potential target for myeloma therapy. As a concept of principle, the present study established a USP5/c-Maf-based luciferase system that was used to screen an FDA-approved drug library. It was found that mebendazole, a typical anthelmintic drug, preferentially induced apoptosis in c-Maf-expressing myeloma cells. Moreover, oral administration of mebendazole delayed the growth of human myeloma xenografts in nude mice but did not show overt toxicity. Further studies showed that the selective antimyeloma activity of mebendazole was associated with the inhibition of the USP5/c-Maf axis. Mebendazole downregulated USP5 expression and disrupted the interaction between USP5 and c-Maf, thus leading to increased levels of c-Maf ubiquitination and subsequent c-Maf degradation. Mebendazole inhibited c-Maf transcriptional activity, as confirmed by both luciferase assays and expression measurements of c-Maf downstream genes. In summary, this study identified mebendazole as a USP5/c-Maf inhibitor that could be developed as a novel antimyeloma agent.

Laparoscopic perigastric mesogastrium excision technique for radical total gastrectomy.

INTRODUCTION: Complete mesogastrium excision (CME) of the perigastric mesogastrium and dissection of lymph nodes (LNs) no. 10 and no. 11 remain technically challenging aspects of laparoscopic radical total gastrectomy (LRTG) plus CME. To address some of these difficulties, we introduced the laparoscopic perigastric mesogastrium excision technique (LPMET) and the concept of the "enjoyable space" to partly modify the procedures of conventional radical surgery and characterize the perigastric space and the surgical plane as well as its boundaries. AIM: To introduce the laparoscopic perigastric mesogastrium excision technique (LPMET) and the "enjoyable space" when undergoing laparoscopic radical total gastrectomy. MATERIAL AND METHODS: From July 2016 to June 2017, 79 cases of upper gastric cancer that were treated by laparoscopic D2 gastrectomy plus CME were investigated. The retrospective database included the patient characteristics, intraoperative and postoperative outcomes, and morbidity and mortality rates depending on the completeness of their medical records. RESULTS: Laparoscopic D2 gastrectomy plus CME was successfully performed in all 79 cases. The mean surgical time was 232.5 ±46.0 min, and the intraoperative blood loss was 67.6 ±52.3 ml. A total of 2245 LNs were retrieved (mean 28.1 ±10.8 retrieved from each specimen). The mean postoperative hospital stay was 10.3 ±1.6 days. The postoperative morbidity rate was 17.7%. After a median follow-up period of 12 months, one patient experienced liver metastasis; of the other 78 patients, none died or experienced tumor recurrence or metastasis. CONCLUSIONS: Laparoscopic perigastric mesogastrium excision technique and the "enjoyable space" could be a novel, minimally invasive approach and space to achieve CME and provide benefit for the dissection of LNs no. 10 and no. 11.

Quantification of enantiomers by mass spectrometry based on chemical derivatization and spectral shape deformation quantitative theory.

A facile mass spectrometric kinetic method for quantitative analysis of chiral compounds was developed by integrating mass spectrometry based on chemical derivatization and the spectral shape deformation quantitative theory. Chemical derivatization was employed to introduce diastereomeric environments to the chiral compounds of interest, resulting in different abundance distribution patterns of fragment ions of the derivatization products of enantiomers in mass spectrometry. The quantitative information of the chiral compounds of interest was extracted from complex mass spectral data by an advanced calibration model derived based on the spectral shape deformation quantitative theory. The performance of the proposed method was tested on the quantitative analysis of R-propranolol in propranolol tablets. Experimental results demonstrated that it could achieve accurate and precise concentration ratio predictions for R-propranolol with an average relative predictive error (ARPE) of about 4%, considerably better than the corresponding results of the mass spectrometric method based on chemical derivatization and the univariate ratiometric model (ARPE: about 12%). The limit of detection (LOD) and limit of quantification (LOQ) of the proposed method for the concentration ratio of R-propranolol were estimated to be 1.5% and 6.0%, respectively. The proposed method is complementary to the existing methods designed for the quantification of enantiomers such as the Cooks kinetic method.

Dichloroacetate enhances the antitumor efficacy of chemotherapeutic agents via inhibiting autophagy in non-small-cell lung cancer.

BACKGROUND: Chemotherapy is still the primary adjuvant strategy of cancer therapy; however, the emergence of multi-drug resistance has been a cause for concern. Autophagy has been demonstrated to have a protective role against chemotherapeutic drugs in cancer cells, and autophagy inhibition is generally considered to be a promising therapeutic strategy. However, the paucity of effective and specific autophagy inhibitors limits its application. PURPOSE: The objective of this study was to explore the effect of DCA, small molecular anti-tumor agent, on the autophagy regulation and chemosensitization in NSCLC cells. METHODS: We investigated the autophagy regulation of dichloroacetate (DCA) by laser confocal microscopy and western blotting in A549 and H1975 cell lines. The MTT assay and flow cytometry was performed for explore the chemosensitization effectiveness of DCA. The results were verified with subcutaneous tumor model in nude mice and the immunohistochemistry was applied for assessing the level of cell apoptosis and autophagy in vivo post treatment. RESULTS: We found that DCA, which exhibited antitumor properties in various carcinoma models, induced apoptosis of non-small cell lung cancer cells (NSCLC) by inhibiting cancer cell autophagy. Furthermore, Perifosine, an AKT inhibitor, can greatly weaken the capacity of inducing apoptosis by DCA. The results indicate that the AKT-mTOR pathway, a main negative regulator of autophagy, is involved in the DCA-induced inhibition of autophagy. Then, we detected the effectiveness of autophagy inhibition by DCA. When used in co-treatment with the chemotherapeutic drug paclitaxel (PTX), DCA markedly decreased cell autophagy, enhanced apoptosis and inhibited proliferation in A549 and H1975 cells. The results of the xenograft experiment demonstrate that co-treatment of PTX and DCA can significantly decrease cell proliferation in vivo and prolong the survival of mice. CONCLUSION: Our results suggest that DCA can inhibit cell autophagy induced by chemotherapeutics, providing a new avenue for cancer chemotherapy sensitization.

Muscone exerts protective roles on alcohol-induced osteonecrosis of the femoral head.

Long-term alcohol abuse causes musculoskeletal disorders, among of which, alcohol-induced osteonecrosis of the femoral head (ONFH) is of concern due to its significant and severe complications. A variety of methods have been attempted to prevent alcohol-induced ONFH, and monomers extracted from Chinese herbs might benefit the disease profoundly. In the current study, muscone, the main ingredient of musk, was used to prevent alcohol-induced ONFH. In vitro, ethanol was used to affect the potential of osteogenesis and proliferation of human bone mesenchymal stem cells (hBMSCs), and beneficial role of muscone was investigated on hBMSCs. In vivo, following the establishment of alcohol-induced ONFH, muscone was employed to treat the diseased rats, which were analyzed by micro-CT scanning and a series of histologic staining. As a result, we found ethanol could significantly suppress osteogenic differentiation of hBMSCs, while muscone held the potential to promote ALP activity and mRNA expressions of COL1 and OCN under ethanol treatment. Meanwhile, imaging analysis revealed muscone could restore BV/TV ratio and bone mineral density of the necrotic femoral head, and the protective role of muscone on alcohol-induced ONFH was further confirmed by histologic examinations. Our study confirmed the protective effect of muscone against alcohol-induced ONFH both in vitro and in vivo. Therefore, muscone may be considered as a valuable therapeutic natural drug for alcohol-induced ONFH in humans.

Saponins from Paris forrestii(Takht.)H.Li displays potent activity against acute myeloid leukemia by suppressing RNF6/AKT/mTOR signaling pathway / 中国药理学与毒理学杂志

Acute myeloid leukemia (AML) is a heterogeneous disease characterized by the accu-mulation of immature myeloid progenitor cells in the bone marrow,compromising of normal hematopoi-esis and ultimately resulting in bone marrow failure. Chemotherapy is the mainstay treatment for all AML patients,however,drug resistance and clinical relapse limits its efficacy.The 5-year survival rate of AML patients is only 26.6%.Survival rates are even lower among patients ages 65 to 74 years (5.3%)and 75 years or older(1.6%).Therefore,exploring novel therapeutic agents is urgent for improving the outcome of patients with AML. Saponins are amphipathic glycosides found in traditional Chinese medicines. In the present study, we isolated a panel of saponins from Paris forrestii (Takht.) H. Li, a unique plant found in Tibet and Yunnan provinces, China. By examining their activities in suppressing acute myeloid leukemia cell proliferation, total saponins from Paris forrestii (TSPf) displayed more potent activity than individual ones.TSPf induced more than 40% AML cell apoptosis within 24 h and decreased the viability of all leukemia cell lines. TSPf-induced apoptosis was confirmed by both Annexin V staining and caspase-3 activation.TSPf downregulated pro-survival proteins Mcl-1,Bcl-xL and Bcl-2,but upreg-ulated the expression of tumor suppressor proteins p53,p27,Bax and Beclin 1.The AKT/mTOR signaling pathway is frequently over activated in various AML cells,and TSPf was found to suppress the activa-tion of both AKT and mTOR,but had no effects on their total protein expression.This was further con-firmed by the inactivation of 4EBP-1 and p70S6K,two typical downstream signal molecules in the AKT/mTOR pathway. More specifically, TSPf-inactivated AKT/mTOR signaling was found to be associated with downregulated RNF6, a recently identified oncogene in AML. RNF6 activated AKT/mTOR, and consistently, knockdown of RNF6 led to inactivation of the AKT/mTOR pathway. Furthermore, TSPf suppressed the growth of AML xenografts in nude mice models. Oral administration of 100 mg·kg-1 body weight almost fully suppressed tumor growth within 14 d, without gross toxicity. This study thus demonstrated that TSPf displays potent anti-AML activity by suppressing the RNF6/AKT/mTOR pathway. Given its low toxicity,TSPf could be developed for the treatment of AML.

A novel STAT3 inhibitor negatively modulates platelet activation and aggregation.

The signal transducer and activator of transcription 3 (STAT3) plays a critical role in platelet functions. This study sought to understand the effects of the STAT3 inhibitor SC99 on platelet activation and aggregation. Immunoblotting assays were applied to measure the effects of SC99 on the STAT3 signaling pathway. A ChronoLog aggregometer was used to evaluate platelet aggregation. A flow cytometer was used to evaluate P-selectin expression in the presence of SC99. AlamarBlue and Annexin-V staining were used to evaluate platelet viability and apoptosis, respectively. A fluorescence microscope was applied to analyze platelet spreading. SC99 inhibited the phosphorylation of JAK2 and STAT3 in human platelets but had no effects on the phosphorylation of AKT, p65 or Src, all of which are involved in platelet activation. Further studies revealed that SC99 inhibited human platelet aggregation induced by collagen and thrombin in a dose-dependent manner. SC99 inhibited thrombin-induced P-selectin expression and fibrinogen binding to single platelets. Moreover, SC99 inhibited platelet spreading on fibrinogen and clot retraction mediated by outside-in signaling. SC99 inhibited platelet aggregation in mice but it did not significantly prolong the bleeding time. Taken together, the present study revealed that SC99 inhibited platelet activation and aggregation as a STAT3 inhibitor. This agent can be developed as a promising treatment for thrombotic disorders.