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.

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.

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.

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.

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.

Association between Th17-related cytokines and risk of non-small cell lung cancer among patients with or without chronic obstructive pulmonary disease.

BACKGROUND: CD4 (+) T helper 17 (Th17) cells play critical roles in inflammation and tumor development. The involvement of Th17 cells in chronic obstructive pulmonary disease (COPD)-type inflammation-associated lung cancer has also been confirmed in animal models. However, to the authors' knowledge, it is unknown whether the role of Th17 cells is different in patients with lung cancer complicated with COPD compared with those without COPD. In the current study, the authors attempted to determine the association between the circulating levels of Th17-related cytokines and the clinical characteristics of non-small cell lung cancer (NSCLC) in patients with or without COPD. METHODS: The authors designed a matched case-control study that included 70 patients with NSCLC with COPD, 148 patients with NSCLC without COPD, and 148 healthy controls. The data regarding the clinicopathological features of these participants were collected. Circulating levels of Th17-related cytokines, including interleukin (IL) 23 (IL-23), IL-17A, IL-17F, IL-22, and tumor necrosis factor-α, were measured. RESULTS: The circulating levels of IL-23, IL-17A, IL-17F, IL-22, and tumor necrosis factor-α were found to be significantly higher in the patients with NSCLC compared with the healthy controls (P<.05). The elevated levels were found to be significantly associated with lung cancer risk (P<.05). However, no significant differences were found between patients with NSCLC with COPD and patients without COPD. It is interesting to note that, among patients with NSCLC without COPD, the levels of these cytokines were consistently higher among patients with stage I to stage IIIA disease compared with those with stage IIIB to stage IV disease (P<.05). In addition, the 5 Th17-related cytokines demonstrated pairwise correlations, with Spearman rank correlation coefficients of 0.646 to 0.888 (P<.05). CONCLUSIONS: The results of the current study indicate a clear association between the Th17-related cytokine profile and the risk of NSCLC complicated by the presence or absence of COPD.

Late surgery for acetabular fractures in a Chinese level I trauma centre: surgical experience and outcomes.

PURPOSE: Delayed surgical management of acetabular fractures, often necessary due to life-threatening concomitant injuries, is a great challenge because delays may potentially increase complications and decrease outcomes. We report clinical outcomes of 61 acetabular fractures treated by delayed open reduction and internal fixation (ORIF) with an injury-to-surgery interval (ISI) of 22-399 days. METHODS: Operations were performed between April 2001 and December 2008. There were 61 cases (42 men 19 women), with an average age of 38 years. All patients were followed for an average of 82 months. Demographic data, fracture pattern, ISI, concomitant injuries, surgical approach, complications and clinical outcomes were recorded and analysed. There were 16 simple fractures (26.2%) and 45 associated fractures (73.8%). Matta criteria were used to evaluate reduction quality. The Merle d'Aubigné and Postel scoring system was employed to assess post-operative functionality. RESULTS: Anatomical reduction was achieved in 45 cases (73.8%). The clinical result was excellent in 38 cases, good in 13, fair in six and poor in four. Osteonecrosis of the femoral head was observed in three cases, and heterotopic ossification was found in 28 cases. Four patients had transient palsy of the sciatic nerve. CONCLUSIONS: ORIF for fresh acetabular fractures might yield a better prognosis; however, for delayed acetabular fractures, clinical outcomes are also predictable when sophisticated surgical techniques are employed. Our results indicate that delayed ORIF could yield satisfactory clinical outcomes in the majority of patients with acetabular fractures.

Subtrochanteric femur fracture treated by intramedullary fixation.

PURPOSE: To discuss surgical technique, operative efficacy and clinical outcome of intramedullary fixation in the treatment of subtrochanteric femur fractures. METHODS: From February 2011 to February 2013, 76 cases of subtrochanteric femur fractures were treated by intramedullary fixation in our hospital, including 53 males and 23 females, with the age range of 37 -72 years (mean 53.5 years). According to Seinsheimer classification, there were 2 cases of type I, 7 type II, 15 type III, 23 type IV and 29 type V. Firstly, all patients underwent closed reduction with the guidance of C-arm fluoroscopy in a traction table. Two cases of type I and 3 cases of type III fractures had ideal closed reduction followed by internal fixation. The others needed additional limited open reduction. Radiographic examination was used to evaluate callus formation and fracture healing in postoperative 1, 3, 6 and 12 months follow-up. Functional recovery was evaluated by Harris Hip Scoring (HHS) system. RESULTS: Patients were followed up for 6-12 months. All fractures were healed except one patient with delayed union. The average bone union time was 4.5 months. According to HHS system, 65 cases were considered as excellent in functional recovery, 8 good, 2 fair and 1 poor. The proportion of the patients with excellent and good recovery was 96.05%. CONCLUSION: Intramedullary fixation is feasible for the treatment of subtrochanteric femur fracture. The accuracy of intraoperative reduction and surgical skill are important for the clinical outcome and the patients' prognosis.

Prognostic significance of VEGF expression in osteosarcoma: a meta-analysis.

Vascular endothelial growth factor (VEGF) is considered as a prime mediator of angiogenesis and has been implicated in carcinogenesis and metastasis. Various studies examined the relationship between VEGF overexpression with the clinical outcome in patients with osteosarcoma but yielded conflicting results. Electronic databases updated to April 2013 were searched to find relevant studies. A meta-analysis was conducted with eligible studies which quantitatively evaluated the relationship between VEGF overexpression and survival of patients with osteosarcoma. Survival data were aggregated and quantitatively analyzed. We performed a meta-analysis of eight studies that evaluated the correlation between VEGF overexpression and survival in patients with osteosarcoma. Combined hazard ratios suggested that VEGF overexpression had an unfavorable impact on overall survival (hazard ratio (HR) = 1.75, 95% confidence interval (CI): 1.21-2.28) in patients with osteosarcoma for overall populations, 2.37 (1.35-3.39) in Asian studies but not in non-Asian studies (HR = 1.51, 95% CI: 0.89-2.14). No significant heterogeneity was observed among all studies. VEGF overexpression indicates a poor prognosis for patients with osteosarcoma. However, the prognostic value of VEGF on survival in osteosarcoma patients still needs further large-scale prospective trials to be clarified.

Adjacent-level symptomatic fracture after percutaneous vertebral augmentation of osteoporotic vertebral compression fracture: a retrospective analysis.

BACKGROUND: Vertebral augmentation is an effective and minimally invasive procedure that is used extensively worldwide for the treatment of osteoporosis vertebral compression fractures (OVCFs). New pain from adjacent vertebra fracture (AVF) after initial cement augmentation has gradually been given attention, but the exact causes of AVF are still controversial. The purpose of this study was to analyze the associated incidence, risk factors, and possible causative mechanism of symptomatic AVF, and to evaluate the intrinsic relationship between cement leakage into the disk and AVF. METHODS: Three hundred and fifty-eight patients (271 women, 87 men; mean age 70.5 ± 9.1 years; range 42-91 years) undergoing vertebral augmentation for their single level of OVCFs were enrolled in the study. Patients were divided into AVF (n = 26) and AVF-free (n = 332) groups, and the groups were compared with respect to preoperative and perioperative parameters, as well as postoperative results. Potential risk factors were evaluated using logistic regression analysis. RESULTS: The rate of symptomatic AVF was 7.3%. The majority of symptomatic AVF, 57.7% of which developed 6 months after the procedures, were located mainly in the thoraco-lumbar vertebra. Significant differences were found between the AVF and AVF-free groups with regard to age, bone mineral density (BMD), and intravertebral clefts (p < 0.05). AVF occurred in six of 28 patients with intravertebral clefts, and five of them developed AVF within 6 months after the procedure. No statistically significant association was observed in the correlation between intradiscal cement leakage and the incidence of symptomatic AVF (p = 0.390). CONCLUSIONS: Older age, lower BMD, and intravertebral clefts are the main risk factors for symptomatic AVF after vertebral augmentation, but intradiscal cement leakage does not increase the risk of AVF. AVF occurs because of the natural progression of osteoporosis. Even distribution of bone cement in the vertebral body is important in OVCF patients with intravertebral clefts.

Mode converter in metal-insulator-metal plasmonic waveguide designed by transformation optics.

A metal-insulator-metal (MIM) waveguide can support two plasmonic modes. Efficient conversion between the two modes can be achieved by reshaping of both phase and power density distributions of the guided mode. The converters are designed with the assistance of transformation optics. We propose two practical configurations for mode conversion, which only consist of homogeneous materials yielded from linear coordinate transformations. The functionalities of the converters are demonstrated by full wave simulations. Without consideration of transmission loss, conversion efficiency of as high as 95% can be realized.

Association of the CTLA-4 +49A/G polymorphism with rheumatoid arthritis in Chinese Han population.

Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) is an important molecule in the regulation of T cells, so the CTLA-4 gene has been considered as a strong candidate associated with T cell-mediated autoimmune diseases such as rheumatoid arthritis (RA). CTLA-4 has many variants and polymorphic forms, among which the +49A/G polymorphism, causing a non-synonymous substitution, has been studied most. However, previous studies of the association between the +49A/G polymorphism of the CTLA-4 gene and RA have provided conflicting results. The aim of this study was to determine the potential relationship of the CTLA-4 +49A/G polymorphism and the risk of RA in Chinese Han population. TaqMan assay was used to genotype the +49A/G polymorphism in 1,489 RA patients and 1,200 healthy controls. Furthermore, a meta-analysis of all studies relating this polymorphism in Chinese population to the risk of RA was performed. The genotype and allele frequencies of the CTLA-4 +49A/G in patients with RA differed significantly from those of controls (P = 0.03 and P = 0.007, respectively). The meta-analysis also revealed that the CTLA-4 +49G allele was associated with an increased risk of RA in Chinese population. Our results suggested that the CTLA-4 gene might contribute to the pathogenesis of RA, and the +49A/G polymorphism of this gene was a risk factor associated with increased RA susceptibility in Chinese Han population.

Nanostructure fabricated by nanosphere lithography assisted with O2 plasma treatment.

Nanosphere lithography is an efficient way to fabricate metallic nanostructures with large area. This paper presents the fabrication of metallic hexagonal nano-pyramid arrays by two dimensional nanospheres lithography assisted with O2 plasma treatment. By O2 plasma treatment, the gap and diameter of nanospheres can be modulated. After electron beam deposition, we can fabricate similar nanostructures with different pyramid gap distances. This method may be an easy way to modulate the geometric parameters of nanostructures.

Optical properties of metal-multi-insulator-metal plasmonic waveguides.

We theoretically study the plasmonic modes in metal-multi-insulator-metal (MMIM) waveguides. Two types of symmetric MMIM structures consisting of three insulators are investigated thoroughly. The effective refractive index, energy confinement, propagation length, and figure of merit are given in terms of geometric parameters. Due to the step index modulation, these properties of MMIM structures differ from the metal-insulator-metal (MIM) structure. Compared with the corresponding MIM structures, MMIM structures can possess either better energy confinement or larger propagation length, which depends on the geometric parameters and the index distribution. Propagation length of up to 10(3) µm and a figure of merit of up to 10(4) are observed for MMIM structure with core thickness of several hundred nanometers.

Minimally invasive percutaneous osteosynthesis for proximal humeral shaft fractures with the PHILOS through the deltopectoral approach.

PURPOSE: The aim of this study was to evaluate clinical outcomes and complications following minimally invasive plate osteosynthesis (MIPO) with the proximal humeral internal locking system (PHILOS) for treating proximal humeral shaft fracture through the deltopectoral approach. METHODS: Between November 2008 and March 2010, 74 patients with unilateral proximal humeral shaft fractures were treated using the MIPO technique with the PHILOS through the deltopectoral approach. Patients received an average follow-up of 16.9 (range, 12-24) months, and the final follow-up included anteroposterior and lateral imaging and recording of postoperative complications. The Constant-Murley shoulder score was used to evaluate function. RESULTS: No intraoperative complications occurred. Postoperative complications included subacromial impingement in four patients. There was no deep infection, neurovascular damage, breakage or implant loosening. All fractures united in an average time of 17.4 (15-25) weeks. In terms of function, the Constant-Murley score was 85.8 points on average (range, 67-100). The range of motion of the involved shoulder was satisfactory, and pain-free in 83.8 % of patients. CONCLUSIONS: Using the MIPO technique with the PHILOS through the deltopectoral approach is a valid and safe method of treating proximal humeral shaft fractures.

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.

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.

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.