BACKGROUND Hepatocellular carcinoma(HCC)is one of the most aggressive tumors worldwide.Chronic inflammation contributes to tumor evolution,and the infiltration of tumor-associated macrophages(TAMs),also known as M2-li...BACKGROUND Hepatocellular carcinoma(HCC)is one of the most aggressive tumors worldwide.Chronic inflammation contributes to tumor evolution,and the infiltration of tumor-associated macrophages(TAMs),also known as M2-like macrophages,is associated with the most aggressive behavior.Therefore,these macrophages provide the primary growth and migratory factors to the tumor cells,including those of HCC.Current therapies are not well optimized for eliminating trans-formed cells or neutralizing the tumor immune microenvironment leukocytes,such as TAMs.Growth differentiation factor 11(GDF11)may represent a promi-sing dual therapeutic target due to its reported anti-tumorigenic and immuno-modulatory properties.AIM To characterize the effects of GDF11 in M2-like macrophages and the HCC cell interaction using a functional in vitro model.METHODS This research used THP-1 and Huh7 cell lines.We applied recombinant GDF11(50 ng/mL)every 24 hours on THP-1 differentiated macrophages with M2-like polarization using interleukin-4 and interleukin-13.Firstly,the GDF11 effects on signaling,viability,proliferation,metabolism,and redox state in macrophages were charac-terized.Subsequently,we extracted conditioned media(CM)from macrophages and performed indirect co-cultures with Huh7 cells.The functional parameters were proliferation and migration assays.Finally,we charac-terized secretion in the CM using the cytokine array membrane assay.RESULTS The present study demonstrated that GDF11 activates the canonical pathway Smad2/3 without cytotoxic or prolif-erative effects.We provide evidence that GDF11 also diminishes the pro-tumoral properties of M2-like macrophages.GDF11 promoted the reduction of the M2-like macrophage marker,cluster of differentiation 206,indicating a loss of pro-tumoral properties in these leukocytes.Furthermore,this molecule induced changes in metabolism and an increase in reactive oxygen species.Using CM derived from GDF11-treated M2-like macrophages,we observed a reduction in the proliferation and migratory capacity of liver cancer cells.Moreover,the cytokine profile was affected by GDF11 stimulus,demonstrating that this molecule alters the pro-tumoral properties of TAMs,which in turn impact the behavior of HCC-derived cells.CONCLUSION This in vitro study suggests that mitigating tumor-promoting or M2-like macrophages with GDF11 may be an effective strategy for controlling the aggressiveness of HCC.展开更多
Background:Understanding how the tumor microenvironment is shaped by various factors is important for the development of new therapeutic strategies.Tumor cells often undergo spontaneous apoptotic cell death in tumor m...Background:Understanding how the tumor microenvironment is shaped by various factors is important for the development of new therapeutic strategies.Tumor cells often undergo spontaneous apoptotic cell death in tumor microen-vironment,these apoptotic cells are histologically co-localized with immunosup-pressive macrophages.However,the mechanism by which tumor cell apoptosis modulates macrophage polarization is not fully understood.In this study,we aimed to explore the tumor promoting effects of apoptotic tumor cells and the signal pathways involved.Methods:Apoptotic cells and macrophages in tumors were detected by immunohistochemical staining.Morphological analysis was performed with Giemsa staining.Lipids generated from apoptotic cells were detected by liq-uid chromatography-mass spectrometry.Phosphatidylserine-containing lipo-somes were prepared to mimic apoptotic cells.The expression of protein was determined by real-time PCR,immunohistochemistry enzyme-linked immunosorbent assay and Western blotting.Mouse malignant ascites and subcu-taneous tumor models were designed for in vivo analysis.Transgenic mice with specific genes knocked out and inhibitors specific to certain proteins were used for the mechanistic studies.Results:The location and the number of apoptotic cells were correlated with that of macrophages in several types of carcinomas.Phosphatidylserine,a lipid molecule generated in apoptotic cells,induced polarization and accumulation of M2-like macrophages in vivo and in vitro.Moreover,sustained administration of phosphoserine promoted tumor growth in the malignant ascites and subcuta-neous tumor models.Further analyses suggested that phosphoserine induced a M2-like phenotype in macrophages,which was related to the activation of phosphoserine receptors including T-cell immunoglobin mucin 4(TIM4)and the FAK-SRC-STAT3 signaling pathway as well as elevated the expression of the histone demethylase Jumonji domain-containing protein 3(JMJD3).Adminis-tration of specific inhibitors of these pathways could reduce tumor progression.Conclusions:This study suggest that apoptotic cell-generated phosphoserine might be a notable signal for immunosuppressive macrophages in tumors,and the related pathways might be potential therapeutic targets for cancer therapy.展开更多
Photothermal therapy(PTT)has received a lot of attention as a promising strategy for eliminating tumors quickly.However,the unavoidable inflammatory response during the treatment might result in a high concentration o...Photothermal therapy(PTT)has received a lot of attention as a promising strategy for eliminating tumors quickly.However,the unavoidable inflammatory response during the treatment might result in a high concentration of M2-like tumor-associated macrophages(TAMs),increasing the risk of tumor recurrence and metastasis.To address this problem,gold-based nanocarriers(PGMP-small interfering RNA(siRNA)nanoparticles(NPs))containing STAT6siRNA,that inhibited M2-like TAM polarization,were designed and investigated for PTT and gene therapy of non-small cell lung cancer(NSCLC).In an NSCLC model,the nanocarriers demonstrated excellent siRNA delivery ability and a high gene transfection rate of up to 90%in macrophages,thus inhibiting the polarization of about 87%of M2-like TAMs and effectively suppressing the invasion and metastasis of NSCLC.Meanwhile,the unique gold nanosphere structure offered improved PTT and contrast-enhanced ultrasound imaging,thus contributing to the efficient elimination and real-time monitoring of the tumor tissues.These nanocarriers with combined gene and photothermal therapeutic capabilities improved the efficacy of single-modality treatment,and showed the potential to inhibit cancer cell recurrence and metastasis to ultimately cure NSCLC.展开更多
Despite significant advances in our understanding of the molecular pathogenesis of liver cancer and the availability of novel pharmacotherapies,liver cancer remains the fourth leading cause of cancer-related mortality...Despite significant advances in our understanding of the molecular pathogenesis of liver cancer and the availability of novel pharmacotherapies,liver cancer remains the fourth leading cause of cancer-related mortality worldwide.Tumor relapse,resistance to current anti-cancer drugs,metastasis,and organ toxicity are the major challenges that prevent considerable improvements in patient survival and quality of life.Calculus bovis(CB),an ancient Chinese medicinal drug,has been used to treat various pathologies,including stroke,convulsion,epilepsy,pain,and cancer.In this editorial,we discuss the research findings recently published by Huang et al on the therapeutic effects of CB in inhibiting the development of liver cancer.Utilizing the comprehensive transcriptomic analyses,in vitro experiments,and in vivo studies,the authors demonstrated that CB treatment inhibits the tumor-promoting M2 phenotype of tumor-associated macrophages via downregulating Wnt pathway.While multiple studies have been performed to explore the molecular mechanisms regulated by CB,this study uniquely shows its role in modulating the M2 phenotype of macrophages present within the tumor microenvironment.This study opens new avenues of future investigations aimed at investigating this drug’s efficacy in various mouse models including the effects of combination therapy,and against drug-resistant tumors.展开更多
目的构建以乙型肝炎病毒核心蛋白(Hepatitis B virus coreprotein,HBc)颗粒为载体的含流感病毒M2基质蛋白胞外功能区(M2 ectodomain,M2e)和核蛋白(Nucleoprotein,NP)保守表位的流感通用疫苗,评价其抗不同亚型流感病毒感染的保护作用。...目的构建以乙型肝炎病毒核心蛋白(Hepatitis B virus coreprotein,HBc)颗粒为载体的含流感病毒M2基质蛋白胞外功能区(M2 ectodomain,M2e)和核蛋白(Nucleoprotein,NP)保守表位的流感通用疫苗,评价其抗不同亚型流感病毒感染的保护作用。方法采用基因工程方法将流感病毒M2e的3拷贝重复片段与NP的CTL表位串联,插入HBc刺突顶端的免疫优势决定区,构建重组表达质粒pET-21a-HBc-3M2e-NP,转化大肠杆菌BL21(DE3),IPTG16℃低温诱导表达。表达的融合蛋白HBc-3M2e-NP经纯化后,电镜观察病毒样颗粒形成情况。纯化的融合蛋白分别经鼻腔和腹腔免疫小鼠,对照组注射等体积的PBS,间接ELISA法检测小鼠血清IgG抗体水平;流式细胞术检测小鼠脾组织中CD4+、CD8+T淋巴细胞水平;以流感病毒攻毒,检测融合蛋白的保护效果。结果重组表达质粒pET-21a-HBc-3M2e-NP经PCR及双酶切鉴定证明构建正确。表达的融合蛋白相对分子质量约为29000,以包涵体和可溶性两种形式表达,且可与鼠抗M2e单抗发生特异性反应。纯化的融合蛋白纯度大于90%,且能够自动装配成病毒样颗粒。用该病毒样颗粒免疫小鼠,可诱导小鼠产生针对不同流感病毒毒株的特异性抗体;2种途径免疫的小鼠脾组织中CD4+T淋巴细胞含量和CD4+/CD8+T淋巴细胞比例均较对照组明显升高;攻毒试验结果显示,具有交叉保护作用。结论构建的流感通用疫苗能够诱导机体产生高水平的体液免疫和细胞免疫应答及有效的交叉保护作用,为流感通用疫苗的深入研究奠定了基础。展开更多
文摘BACKGROUND Hepatocellular carcinoma(HCC)is one of the most aggressive tumors worldwide.Chronic inflammation contributes to tumor evolution,and the infiltration of tumor-associated macrophages(TAMs),also known as M2-like macrophages,is associated with the most aggressive behavior.Therefore,these macrophages provide the primary growth and migratory factors to the tumor cells,including those of HCC.Current therapies are not well optimized for eliminating trans-formed cells or neutralizing the tumor immune microenvironment leukocytes,such as TAMs.Growth differentiation factor 11(GDF11)may represent a promi-sing dual therapeutic target due to its reported anti-tumorigenic and immuno-modulatory properties.AIM To characterize the effects of GDF11 in M2-like macrophages and the HCC cell interaction using a functional in vitro model.METHODS This research used THP-1 and Huh7 cell lines.We applied recombinant GDF11(50 ng/mL)every 24 hours on THP-1 differentiated macrophages with M2-like polarization using interleukin-4 and interleukin-13.Firstly,the GDF11 effects on signaling,viability,proliferation,metabolism,and redox state in macrophages were charac-terized.Subsequently,we extracted conditioned media(CM)from macrophages and performed indirect co-cultures with Huh7 cells.The functional parameters were proliferation and migration assays.Finally,we charac-terized secretion in the CM using the cytokine array membrane assay.RESULTS The present study demonstrated that GDF11 activates the canonical pathway Smad2/3 without cytotoxic or prolif-erative effects.We provide evidence that GDF11 also diminishes the pro-tumoral properties of M2-like macrophages.GDF11 promoted the reduction of the M2-like macrophage marker,cluster of differentiation 206,indicating a loss of pro-tumoral properties in these leukocytes.Furthermore,this molecule induced changes in metabolism and an increase in reactive oxygen species.Using CM derived from GDF11-treated M2-like macrophages,we observed a reduction in the proliferation and migratory capacity of liver cancer cells.Moreover,the cytokine profile was affected by GDF11 stimulus,demonstrating that this molecule alters the pro-tumoral properties of TAMs,which in turn impact the behavior of HCC-derived cells.CONCLUSION This in vitro study suggests that mitigating tumor-promoting or M2-like macrophages with GDF11 may be an effective strategy for controlling the aggressiveness of HCC.
基金NationalNatural Science Foundation of China:National Science Foundation for ExcellentYoung Scholars,Grant/Award Number:32122052NationalNatural Sci-ence Foundation of China:NationalNat-ural Science FoundationRegional Inno-vation and Development,Grant/Award Number:U19A2003NationalNatural Science Foundation ofChina:National Science Foundation forYoung Scholars,Grant/Award Number:81902662。
文摘Background:Understanding how the tumor microenvironment is shaped by various factors is important for the development of new therapeutic strategies.Tumor cells often undergo spontaneous apoptotic cell death in tumor microen-vironment,these apoptotic cells are histologically co-localized with immunosup-pressive macrophages.However,the mechanism by which tumor cell apoptosis modulates macrophage polarization is not fully understood.In this study,we aimed to explore the tumor promoting effects of apoptotic tumor cells and the signal pathways involved.Methods:Apoptotic cells and macrophages in tumors were detected by immunohistochemical staining.Morphological analysis was performed with Giemsa staining.Lipids generated from apoptotic cells were detected by liq-uid chromatography-mass spectrometry.Phosphatidylserine-containing lipo-somes were prepared to mimic apoptotic cells.The expression of protein was determined by real-time PCR,immunohistochemistry enzyme-linked immunosorbent assay and Western blotting.Mouse malignant ascites and subcu-taneous tumor models were designed for in vivo analysis.Transgenic mice with specific genes knocked out and inhibitors specific to certain proteins were used for the mechanistic studies.Results:The location and the number of apoptotic cells were correlated with that of macrophages in several types of carcinomas.Phosphatidylserine,a lipid molecule generated in apoptotic cells,induced polarization and accumulation of M2-like macrophages in vivo and in vitro.Moreover,sustained administration of phosphoserine promoted tumor growth in the malignant ascites and subcuta-neous tumor models.Further analyses suggested that phosphoserine induced a M2-like phenotype in macrophages,which was related to the activation of phosphoserine receptors including T-cell immunoglobin mucin 4(TIM4)and the FAK-SRC-STAT3 signaling pathway as well as elevated the expression of the histone demethylase Jumonji domain-containing protein 3(JMJD3).Adminis-tration of specific inhibitors of these pathways could reduce tumor progression.Conclusions:This study suggest that apoptotic cell-generated phosphoserine might be a notable signal for immunosuppressive macrophages in tumors,and the related pathways might be potential therapeutic targets for cancer therapy.
基金This work was supported by the Natural Science Foundation of China(Nos.81873898,81960316,81871411,32011530115,and 32025021)the Science&Technology Bureau of Ningbo City(Nos.2020Z094 and 2021Z072)+1 种基金the Excellent Member of Youth Innovation Promotion Association Foundation of CAS(No.Y2021079)the Ningbo 3315 Innovative Teams Program,China(No.2019A-14-C).
文摘Photothermal therapy(PTT)has received a lot of attention as a promising strategy for eliminating tumors quickly.However,the unavoidable inflammatory response during the treatment might result in a high concentration of M2-like tumor-associated macrophages(TAMs),increasing the risk of tumor recurrence and metastasis.To address this problem,gold-based nanocarriers(PGMP-small interfering RNA(siRNA)nanoparticles(NPs))containing STAT6siRNA,that inhibited M2-like TAM polarization,were designed and investigated for PTT and gene therapy of non-small cell lung cancer(NSCLC).In an NSCLC model,the nanocarriers demonstrated excellent siRNA delivery ability and a high gene transfection rate of up to 90%in macrophages,thus inhibiting the polarization of about 87%of M2-like TAMs and effectively suppressing the invasion and metastasis of NSCLC.Meanwhile,the unique gold nanosphere structure offered improved PTT and contrast-enhanced ultrasound imaging,thus contributing to the efficient elimination and real-time monitoring of the tumor tissues.These nanocarriers with combined gene and photothermal therapeutic capabilities improved the efficacy of single-modality treatment,and showed the potential to inhibit cancer cell recurrence and metastasis to ultimately cure NSCLC.
基金Supported by the National Institutes of Health grants,No.K99HL146954 and No.R00HL146954the UTHSC College of Pharmacy Research Seed Grant award,No.2023.
文摘Despite significant advances in our understanding of the molecular pathogenesis of liver cancer and the availability of novel pharmacotherapies,liver cancer remains the fourth leading cause of cancer-related mortality worldwide.Tumor relapse,resistance to current anti-cancer drugs,metastasis,and organ toxicity are the major challenges that prevent considerable improvements in patient survival and quality of life.Calculus bovis(CB),an ancient Chinese medicinal drug,has been used to treat various pathologies,including stroke,convulsion,epilepsy,pain,and cancer.In this editorial,we discuss the research findings recently published by Huang et al on the therapeutic effects of CB in inhibiting the development of liver cancer.Utilizing the comprehensive transcriptomic analyses,in vitro experiments,and in vivo studies,the authors demonstrated that CB treatment inhibits the tumor-promoting M2 phenotype of tumor-associated macrophages via downregulating Wnt pathway.While multiple studies have been performed to explore the molecular mechanisms regulated by CB,this study uniquely shows its role in modulating the M2 phenotype of macrophages present within the tumor microenvironment.This study opens new avenues of future investigations aimed at investigating this drug’s efficacy in various mouse models including the effects of combination therapy,and against drug-resistant tumors.
基金Supported by Chongqing Innovation and Entrepreneurship Program for Returned Overseas Scholars(No.RC2020-2)Chongqing Natural Science Foundation(No.cstc2018jcyjAX0120,cstc2020jcyj-msxm X0007)National Natural Science Foundation of China(No.31801139)。
文摘目的构建以乙型肝炎病毒核心蛋白(Hepatitis B virus coreprotein,HBc)颗粒为载体的含流感病毒M2基质蛋白胞外功能区(M2 ectodomain,M2e)和核蛋白(Nucleoprotein,NP)保守表位的流感通用疫苗,评价其抗不同亚型流感病毒感染的保护作用。方法采用基因工程方法将流感病毒M2e的3拷贝重复片段与NP的CTL表位串联,插入HBc刺突顶端的免疫优势决定区,构建重组表达质粒pET-21a-HBc-3M2e-NP,转化大肠杆菌BL21(DE3),IPTG16℃低温诱导表达。表达的融合蛋白HBc-3M2e-NP经纯化后,电镜观察病毒样颗粒形成情况。纯化的融合蛋白分别经鼻腔和腹腔免疫小鼠,对照组注射等体积的PBS,间接ELISA法检测小鼠血清IgG抗体水平;流式细胞术检测小鼠脾组织中CD4+、CD8+T淋巴细胞水平;以流感病毒攻毒,检测融合蛋白的保护效果。结果重组表达质粒pET-21a-HBc-3M2e-NP经PCR及双酶切鉴定证明构建正确。表达的融合蛋白相对分子质量约为29000,以包涵体和可溶性两种形式表达,且可与鼠抗M2e单抗发生特异性反应。纯化的融合蛋白纯度大于90%,且能够自动装配成病毒样颗粒。用该病毒样颗粒免疫小鼠,可诱导小鼠产生针对不同流感病毒毒株的特异性抗体;2种途径免疫的小鼠脾组织中CD4+T淋巴细胞含量和CD4+/CD8+T淋巴细胞比例均较对照组明显升高;攻毒试验结果显示,具有交叉保护作用。结论构建的流感通用疫苗能够诱导机体产生高水平的体液免疫和细胞免疫应答及有效的交叉保护作用,为流感通用疫苗的深入研究奠定了基础。
