The study aimed to investigate the protective effect of tanshinone IIA against cardiac hypertrophy in spontaneously hypertensive rats (SHRs) through the Cys-C/Wnt signaling pathway. Thirty SHRs were randomly divided into cardiac hypertrophy, low- and high-dose tanshinone IIA groups. Ten Wistar-Kyoto rats were selected as control group. The systolic blood pressure (SBP), heart weight (HW), left ventricular weight (LVW) and body weight (BW) of all rats were recorded. HE staining and qRT-PCR were applied to observe the morphology of myocardial tissue and mRNA expressions of COL1A1 and COL3A1. ELISA and Western blotting were used to measure the serum asymmetric dimethylarginine (ADMA), nitric oxide (NO) and cardiac troponin I (cTnI) levels, and the expressions of the Cys-C/Wnt signaling pathway-related proteins, eNOS and Nox4. Compared with the cardiac hypertrophy group, the SBP, HW/BW, LVW/BW, swelling degree of myocardial cells, COL1A1 and COL3A1 mRNA expressions, serum cTnI and ADMA levels, and the Cys-C/Wnt signaling pathway-related proteins and Nox4 expressions in the low- and high-dose tanshinone IIA groups were decreased, but the endothelial NO synthase (eNOS), phosphorylated eNOS (Ser1177) and NO expressions were increased. No significant difference was found between the low- and high-dose tanshinone IIA groups. Our study indicated a protective effect of tanshinone IIA against cardiac hypertrophy in SHRs through inhibiting the Cys-C/Wnt signaling pathway.

Figure S1 shows cleaved caspase 3 staining and quantification in CD47 WT and CD47 KO xenografts.

Figure S4 shows Sanger sequencing of GL261 clone 29.

Supplementary Table 1. Primers used for qRT-PCR Supplementary Fig. 1. Heterozygous IDH1R132H/WT enhanced therapy-induced senescence in glioma cells. Supplementary Fig. 2. a. Schematic graph of genome editing strategy with two guide RNAs to knock out human ATRX. b. The ATRX gene sequence of two clones (C4, C10) after genome editing with CRISPR/Cas9 and two ARTX guide RNAs. c. Transwell assays of cell migration showing ATRX knockout in mIDH1 WT cells decreased cell migration. Supplementary Fig. 3. ATRX knockout in SVG IDH1 WT cells did not affect cell growth and migration. Supplementary Fig. 4. a, b. BrdU incorporation assays. c. Cell cycle analysis with PI and flow cytometry. Cells were synchronized by serum withdrawing for 24 h, followed by serum replenishing for 18 h. d. Western blot analyses of Y-H2AX and DNA damage response pathways in cells. Quantification of protein level changes was from the average of three independent experiments. Supplementary Fig. 5. IDH1R132H/WT and ATRX knockout did not induce ALT and alter chromosome numbers in SVG astroglial cells. Supplementary Fig. 6. Effect of AGI-5198 (a) and 5-Aza (b) on SASP expression. Supplementary Fig. 7. M0 human primary macrophages responses to LPS and IL4. Supplementary Fig. 8. ABT-263 decreased SA-beta-gal staining.

TanshinoneⅡA (TanⅡA) is a noteworthy lipophilic diterpene compound derived from the dried roots of the Traditional Chinese Medicine Danshen () that has various pharmacological properties, including anti-inflammatory, antibacterial, and antioxidative effects. Sepsis is a life-threatening organ dysfunction induced by a dysregulated host response to infection. Recently, increasing attention has been paid to sepsis-induced dysfunction of the intestine, car-diovascular system, lungs, kidneys, liver, and other organs. Experimental studies have shown that TanⅡA has therapeutic potential for sepsis-induced organ dysfunction owing to its anti-inflammatory, anti-apoptotic and regulatory effects on multiple signalling pathways. The purpose of this article is to evaluate the potential multiorgan protective effects of TanⅡA in sepsis.

Sepsis is a life-threatening organ dysfunction caused by an abnormal infection-induced immune response. Despite significant advances in supportive care, sepsis remains a considerable therapeutic challenge and is the leading cause of death in the intensive care unit (ICU). Sepsis is characterized by initial hyper-inflammation and late immunosuppression. Therefore, immune-modulatory therapies have great potential for novel sepsis therapies. Ubiquitination is an essential post-translational protein modification, which has been known to be intimately involved in innate and adaptive immune responses. Several E3 ubiquitin ligases have been implicated in innate immune signaling and T-cell activation and differentiation. In this article, we review the current literature and discuss the role of E3 ligases in the regulation of immune response and their effects on the course of sepsis to provide insights into the prevention and therapy for sepsis.

Figure S5 shows Arg-1 staining and quantification in xenografts from CD47 WT +/- TNC KD, and CD KO +/- TNC KD.

Figure S3 shows INOS and TGM2 staining and quantification in CD47 WT and CD47 KO xenografts.

Figure S2 shows Iba-1 staining in CD47 WT and CD47 KO xenografts.

Abstract Human stem‐cell‐derived extracellular vesicles (EVs) are currently being investigated for cell‐free therapy in regenerative medicine applications, but the lack of noninvasive imaging methods to track EV homing and uptake in injured tissues has limited the refinement and optimization of the approach. Here, we developed a new labelling strategy to prepare magnetic EVs (magneto‐EVs) allowing sensitive yet specific MRI tracking of systemically injected therapeutic EVs. This new labelling strategy relies on the use of ‘sticky’ magnetic particles, namely superparamagnetic iron oxide (SPIO) nanoparticles coated with polyhistidine tags, to efficiently separate magneto‐EVs from unencapsulated SPIO particles. Using this method, we prepared pluripotent stem cell (iPSC)‐derived magneto‐EVs and subsequently used MRI to track their homing in different animal models of kidney injury and myocardial ischemia. Our results showed that iPSC‐derived EVs preferentially accumulated in the injury sites and conferred substantial protection. Our study paves a new pathway for preparing highly purified magnetic EVs and tracking them using MRI towards optimized, systemically administered EV‐based cell‐free therapies.

The coronavirus disease 2019 (COVID-19) pandemic has become a public health emergency of global concern. In China, traditional Chinese medicine has been widely administered to COVID-19 patients without sufficient evidence. To evaluate the efficacy of Shenhuang Granule (SHG) for treating critically ill patients with COVID-19, we included in this study 118 patients who were admitted to the ICU of Tongji Hospital between January 28, 2020 and March 28, 2020. Among these patients, 33 (27.9%) received standard care plus SHG (treatment group) and 85 (72.1%) received standard care alone (control group). Enrolled patients had a median (IQR) age of 68 (57-75) years, and most (79 [67.1%]) were men. At end point of this study, 83 (70.3%) had died in ICU, 29 (24.5%) had been discharged from ICU, and 6 patients (5.2%) were still in ICU. Compared with control group, mortality was significantly lower in treatment group (45.4% vs. 80%, p < .001). Patients in treatment group were less likely to develop acute respiratory distress syndrome (ARDS) (12 [36.3%] vs. 54 [63.5%], p = 0.012) and cardiac injury (5 [15.1%] vs. 32 [37.6%], p = 0.026), and less likely to receive mechanical ventilation (22 [66.7%] vs. 72 [84.7%], p = 0.028) than those in control group. The median time from ICU admission to discharge was shorter in treatment group (32 [20?73] days vs. 76 [63?79] days, p = 0.0074). These findings suggest that SHG treatment as a complementary therapy might be effective for critically ill adults with COVID-19 and warrant further clinical trials.

Tumor-associated microglia/macrophages (TAMs) are the main innate immune effector cells in malignant gliomas and have both pro- and anti-tumor functions. The plasticity of TAMs is partially dictated by oncogenic mutations in tumor cells. Heterozygous IDH1 mutation is a cancer driver gene prevalent in grade II/III gliomas, and IDH1 mutant gliomas have relatively favorable clinical outcomes. It is largely unknown how IDH mutation alters TAM phenotypes to influence glioma growth. Here we established clinically relevant isogenic glioma models carrying monoallelic IDH1 R132H mutation (IDH1R132H/WT) and found that IDH1R132H/WT significantly downregulated immune response-related pathways in glioma cells, indicating an immunomodulation role of mutant IDH1. Co-culturing IDH1R132H/WT glioma cells with human macrophages promoted anti-tumor phenotypes of macrophages and increased macrophage migration and phagocytic capacity. In orthotopic xenografts, IDH1R132H/WT decreased tumor growth and prolonged animal survival, accompanied by increased TAM recruitment and upregulated phagocytosis markers, suggesting the induction of anti-tumor TAM functions. Using human cytokine arrays that query 36 proteins, we identified significant downregulation of ICAM-1/CD54 in IDH1R132H/WT gliomas, which was further confirmed by ELISA and immunoblotting analyses. ICAM1 gain-offunction studies revealed that ICAM1 downregulation in IDH1R132H/WT cells played a mechanistic role to mediate the immunomodulation function of IDH1R132H/WT. ICAM-1 silencing in IDH1 wild-type glioma cells decreased tumor growth and increased the anti-tumor function of TAMs. Together, our studies support a new TAM-mediated phagocytic function within IDH1 mutant gliomas, and improved understanding of this process may uncover novel approaches to targeting IDH1 wild type gliomas.

目的:探讨丹参酮ⅡA对心肌成纤维细胞损伤的保护作用及可能机制.方法:分离和培养大鼠心肌成纤维细胞(Cardiac Fibroblast,CFs).以脂多糖(LPS)刺激CFs建立脓毒症心肌损伤体外模型,以不同浓度丹参酮ⅡA预处理CFs 30 min后,给予LPS刺激,设对照组,LPS模型组,LPS+不同浓度TSA(2μmol/L、10μmol/L、50μmol/L)组;采用蛋白质免疫印迹试验法(Western blotting)检测CFs的NLRP3和Caspase-1蛋白表达水平,酶联免疫吸附试验(ELISA)检测细胞上清中IL-1β和IL-18的含量.结果:LPS刺激24 h后,CFs的NLRP3蛋白的表达水平最高.LPS模型组NLRP3和Caspase-1蛋白的表达较对照组明显升高(P<0.01).与LPS模型组比较,丹参酮ⅡA(10μmol/L、50μmol/L)处理组NLRP3和Caspase-1蛋白的表达均明显降低(P<0.05);丹参酮ⅡA处理组的IL-1β和IL-18水平均明显低于LPS模型组(P<0.05).结论:丹参酮ⅡA能抑制LPS诱导的心肌成纤维细胞内NLRP3/Caspase-1炎症反应信号通路分子的表达,这可能是其保护心肌细胞的分子机制之一.

BACKGROUND:Coronavirus disease 2019 (COVID-19) is still a pandemic, with a high mortality rate in severe/critical cases. Therapies based on the Shenghuang Granule have proved helpful in viral infection and septic shock.HYPOTHESIS/PURPOSE:The objective of the current study was to compare the efficacy and safety of the traditional Chinese medicine, Shenhuang Granule, with standard care in hospitalized patients with severe/critical COVID-19.STUDY DESIGN AND METHODS:This was an open-label, multicenter, randomized, controlled clinical trial. At 4 medical centers, a total of 111 severe/critical patients were randomly assigned to receive Shenhuang Granule (SHG group) twice a day for 14 days, in addition to standard care, or to receive standard care alone (Control group). The maximal follow up time was 75 days. The clinical endpoint was clinical improvement and mortality.RESULTS:54 patients were assigned to the control group and 57 to the SHG group. The overall mortality was 75.9% (41/54) in the control group, and 38.6% (22/57) in the SHG group (p < 0.01 vs. control). The post hoc analysis showed that in the severe category, the mortality of the control group vs. the SHG group was 58.8% (10/17) vs. 5.3% (1/19) (p < 0.01); while in the critical category, it was 83.8% (31/37) vs. 55.3% (21/38) (p < 0.05). In the severe category, the mortality of patients who eventually received an invasive ventilator in the control vs. the SHG group was 58.8% (10/17) vs. 0 (0/19) (p < 0.01). Administration of SHG was associated with increased lymphocytes and decreased adverse events.CONCLUSION:Shenhuang Granule is a promising integrative therapy for severe and critical COVID-19.

Abstract Heterozygous isocitrate dehydrogenase (IDH) R132H mutation (IDH1R132H/WT) is an early event during gliomagenesis. Clinically, patients with glioma carrying mutant IDH1 respond better to antitumor therapies. However, the mechanism by which IDH1 mutations contribute to gliomagenesis and therapeutic response remains elusive. Here we report that senescence is involved in the improved therapeutic responses of mutant IDH1 glioma cells. Knocking-in IDH1R132H/WT in glioma cells significantly enhanced gliomas cell senescence in response to temozolomide and radiation via a DNA-damage mediated mechanism. We further asked if senescence plays a role in IDH1R132H/WT-induced gliomagenesis. Together with ATRX knockout and p53/RB loss, IDH1R132H/WT transformed nonneoplastic human astroglial cells to form tumors in mouse brains. In-depth characterization revealed that a subset of these precancerous cells underwent senescence-like phenotypic changes, including flat and enlarged-cell morphology, increased senescence marker expression, decreased cell proliferation, and cell-cycle arrest at the G2–M phase. Mechanistic studies indicated that the combination of glioma driver genes (p53/RB/IDH1/ATRX) dramatically increased DNA damage and activated DNAdamage response (DDR) pathways ATR/ATR and Chk1/Chk2 in senescent cells. To determine how senescent cells drive tumor formation, we investigated non–cell-autonomous mechanisms such as senescence-associated secretory phenotype (SASP), a panel of proinflammatory and tissue-remodeling factors implicated in a tumor-permissive microenvironment. We found that astroglial cells carrying p53/RB/ATRX loss and IDH1R132H/WT upregulated key factors in SASP via an epigenetic-mediated mechanism. Our work suggests that drugs that specifically eliminate senescent cells could help kill precancerous cells and senescent tumor cells following antitumor therapies. Implications: The mechanisms by which IDH1 mutations contribute to gliomagenesis and therapeutic responses remain incompletely characterized; this work reveals senescence as a novel mechanism of IDH-mutant–mediated biological impact and describes new therapeutic opportunities concerning IDH1-mutant gliomas.

Glioblastoma (GBM, WHO grade IV glioma) is the most common and lethal malignant brain tumor in adults with a dismal prognosis. The extracellular matrix (ECM) supports GBM progression by promoting tumor cell proliferation, migration, and immune escape. Uridine diphosphate (UDP)-glucose 6-dehydrogenase (UGDH) is the rate-limiting enzyme that catalyzes the biosynthesis of glycosaminoglycans that are the principal component of the CNS ECM. We investigated how targeting UGDH in GBM influences the GBM immune microenvironment, including tumor-associated microglia/macrophages (TAMs) and T cells. TAMs are the main immune effector cells in GBM and can directly target tumor cells if properly activated. In co-cultures of GBM cells and human primary macrophages, UGDH knockdown in GBM cells promoted macrophage phagocytosis and M1-like polarization. In orthotropic human GBM xenografts and syngeneic mouse glioma models, targeting UGDH decreased ECM deposition, increased TAM phagocytosis marker expression, reduced M2-like TAMs and inhibited tumor growth. UGDH knockdown in GBM cells also promoted cytotoxic T cell infiltration and activation in orthotopic syngeneic mouse glioma models. The potent and in-human-use small molecule GAG synthesis inhibitor 4-methylumbelliferone (4-MU) was found to inhibit GBM cell proliferation and migration in vitro, mimic the macrophage and T-cell responses to UGDH knockdown in vitro and in vivo and inhibit growth of orthotopic murine GBM. Our study shows that UGDH supports GBM growth through multiple mechanisms and supports the development of ECM-based therapeutic strategies to simultaneously target tumor cells and their microenvironment.

目的 观察丹参酮ⅡA磺酸钠对大鼠肠缺血再灌注(Ⅱ/R)肺损伤的作用及影响机制.方法 2019年1—12月于华中科技大学同济医学院附属同济医院进行实验.将30只健康雄性SD大鼠随机数字表法分为假手术+NS组(Sham组),肠缺血再灌注+NS组(Ⅱ/R组),肠缺血再灌注+丹参酮ⅡA磺酸钠注射液组(TanshinoneⅡA组),每组10只.实验结束后处死大鼠取肺组织及支气管肺泡灌洗液(BALF)标本,观察大鼠肺组织病理学改变;测定各组大鼠肺组织湿/干比(W/D);检测大鼠动脉血PaO2,BALF蛋白浓度及炎性细胞因子IL-1β、IL-6及TNF-α水平;检测大鼠肺组织TLR4及pNF-κB p65蛋白表达.结果 肺组织损伤评分、W/D值比较,Sham组<TanshinoneⅡA组<Ⅱ/R组(F/P=178.368/0.000、223.028/0.000);动脉血PaO2比较,Sham组>TanshinoneⅡA组>Ⅱ/R组(F/P=190.446/0.000);BALF蛋白浓度及BALF炎性细胞因子IL-1β、IL-6及TNF-α水平比较,Sham组<TanshinoneⅡA组<Ⅱ/R组(F/P=450.405/0.000、234.850/0.000、1679.317/0.000、543.886/0.000);肺组织TLR4及pNF-κB p65蛋白表达比较,Sham组<TanshinoneⅡA组<Ⅱ/R组(F/P=1434.247/0.000、334.075/0.000).结论 丹参酮ⅡA磺酸钠对Ⅱ/R所致肺损伤具有明显的保护作用,其保护作用可能与抑制TLR4/NF-κB的活化进而抑制肺部炎性反应有关.

目的:探讨PARP-1抑制剂3-AB对肝癌细胞系MHCC97-H和SMMC7721及正常肝细胞系L02的增殖与凋亡的影响.方法:细胞增殖试验观察不同浓度3-AB对三种不同细胞系细胞的增殖作用.AnnexinV荧光探针标记,流式细胞学检查观察不同浓度3-AB对不同细胞系细胞凋亡的影响.结果:当3-AB浓度分别为5mM、10mM与20 mM时,与对照组(0mM)相比,在培养第6天时开始出现增殖明显减慢,出现统计学差异(p＜0.05),第九天差异明显(p＜0.05).随着浓度增加,其对肿瘤细胞系MHCC97-H和SMMC7721细胞增殖的抑制程度增加,细胞数均逐渐减少;而同样浓度梯度3-AB对人类肝细胞系L02生长则无明显的抑制作用.进一步实验发现,当3-AB浓度为5mM、10mM与20 mM时,均可诱导肝癌细胞株MHCC97-H和SMMC7721凋亡,与对照组(0mM)比较均有统计学差异(p＜0.05),且细胞凋亡率与3-AB的药物浓度相关:浓度越高,凋亡越明显.而同等浓度3-AB对肝脏细胞系L02无明显的促进凋亡作用.结论:3-AB可以抑制肝癌肿瘤细胞的增殖,促进肿瘤细胞的凋亡,对正常肝脏细胞无明显毒害作用,具有治疗肝癌的的潜在应用价值.

目的 研究在裸鼠原位种植人肝癌模型中,下调N-cadherin表达对肝癌转移能力的影响.方法 通过原位种植的方法,将极弱表达N-cadherin的LM3-N7细胞与正常表达N-cadherin的LM3-E2细胞分别注射到两组BALB/c裸鼠的肝脏内,6周后取裸鼠肺、肝内转移灶,通过苏木精-伊红(HE)染色观察肺、肝转移灶的情况,分析N-cadherin表达的下调对肝癌细胞转移能力的影响.结果 解剖裸鼠,观察腹水形成及转移灶发现,分别注射LM3-N7和LM3-E2细胞克隆的裸鼠腹水形成(腹腔转移)、肝转移及肺内转移的发生率分别为46.2％(6/13)v s.0％(0/12),46.2％(6/13)vs.41.7％(5/12),23.1％(3/13)vs.16.7％(2/12),统计学分析显示,在腹水形成方面两组差异具有统计学意义(P=0.026),肺转移及肝内转移发生率的差异无统计学意义(均P>0.05).结论 下调N-cadherin表达对裸鼠原位移植癌转移(主要是腹腔转移)具有一定的促进作用,为N-cadherin在肝癌转移机制方面的下一步研究奠定了一定理论基础.