Caffeine modulates the malignant biological behaviors of human brain glioma U-373MG cells by regulating the FAK/AKT/ROCK pathway / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：探讨咖啡因通过调节FAK/AKT/ROCK信号通路来影响人脑胶质瘤U-373MG细胞的增殖、迁移和侵袭能力。方法：常规培养U-373MG细胞，将其分为对照组、咖啡因低剂量（1 mmol/L）组、咖啡因高剂量（2 mmol/L）组、PF573228组（FAK抑制剂，1 μmol/L）、咖啡因高剂量+SC79组（AKT激活剂，8 mg/L）。用CCK-8法、Transwell小室实验、流式细胞术和WB法分别检测各组U-373MG细胞的增殖、迁移、侵袭及凋亡，以及U-373MG细胞中p-FAK、p-AKT、p-ROCK、Ki67、MMP-9蛋白表达水平。建立U-373MG细胞裸鼠移植瘤模型，观察咖啡因对移植瘤生长的影响，WB法检测移植瘤组织中相关蛋白的表达。结果：咖啡因、PF573228可显著抑制U-373MG细胞的增殖、迁移、侵袭能力，促进U-373MG细胞凋亡，抑制p-FAK、p-AKT、p-ROCK、Ki67、MMP-9蛋白的表达（均P<0.05），SC79则可部分逆转咖啡因对U-373MG细胞的作用（均P<0.05）。咖啡因可显著抑制移植瘤的生长及移植瘤组织中上述相关蛋白的表达（均P<0.05）。结论：咖啡因可通过抑制FAK/AKT/ROCK信号通路抑制U-373MG细胞的增殖、迁移和侵袭能力，并促进其凋亡。

PIEZO1 Channel is Involved in Electric Field Guided Cell Migration / 生物化学与生物物理进展

ObjectiveDisruption of epithelial layer may instantaneously induce the generation of endogenous electric fields, which was proved to play an important role in guiding the cell migration and promoting wound healing. PIEZO1 is a kind of mechanic sensitive channel, may be regulated by voltage, is proved to involve in chemotactic migration of cells and play an important role in the process of wound healing. In this paper, the role of PIEZO1 and its downstream proteins FAK and integrin β1 in the electric field guided cell migration were investigated by HaCaT cells (human immortalized keratinocyte). MethodsCell migration was tracked by Living Cell Imaging System in directed current (DC) electric field (EF). Inhibitors and RNAi techniques were applied to study the function of PIEZO1 and other related proteins in electric fields. Western blot was used to detect the expression and phosphorylation levels of integrin β1 and FAK in electric field guided migration under EF stimulation. ResultsPiezo1 RNAi as well as Ruthenium red and GsMTx4 treatment all significantly inhibited the electrotaxis of HaCaT cells. Electric field stimulation with GsMTx4 treatment alone increased FAK phosphorylation level and the expression of integrin β1. Electric field promoted the expression level of integrin β1 and the phosphorylation level of FAK. Inhibiting the expression of PIEZO1 by RNAi significantly attenuated the phosphorylation level of FAK under EF stimulation. Inhibition of integrin β1 and FAK by inhibitor significantly decrease the electric field guided cell migration. ConclusionPIEZO1 as well as integrin β1 and FAK are involved in the electric field guided cell migration of HaCaT cells. Electric field signals regulate the expression of integrin β1 and the activation of FAK through PIEZO1-mediated signal pathway to orchestrate cell migration.

nteraction between FAK/αB-crystalline is important for viability of the glioblastoma cells

Abstract Glioblastoma multiforme is a tumor of the central nervous system. Focal Adhesion Kinase (FAK) and αB-crystalline are two proteins involved in glioblastoma development. In this study, we investigated whether the FAK/αB-crystalline interaction is important for glioblastoma cells, we aimed to investigate the interaction of these two proteins in the glioblastoma multiforme cell line U87-MG. Two peptides named FP01 peptide (derived from αB-crystalline) and FP02 peptide (derived from FAK) were synthesized for this study. Treatment of U87-MG with the peptides FP01 and FP02 in the concentration at 50 µM reduced the viability cellular to around 41% and 51%, respectively. Morphological alterations in the cells treated with the peptides when compared to the control were observed. This study suggests that the interaction between FAK and αB-crystalline is important for the viability of glioblastoma cells

DCLK1 Promotes Malignancy of A549 Cell Line by Activating FAK/PI3K/AKT/mTOR Pathway / 肿瘤防治研究

Objective To investigate the effects of doublecortin-like kinase 1 (DCLK1) on the malignant biological behaviors, such as proliferation, migration, and invasion, of A549 cell line and their corresponding mechanisms. Methods DCLK1-overexpressing A549 cell lines were established through lentiviral infection, and DCLK1 expression was validated by using RT-PCR and Western blot analysis. Proliferation ability was assessed with CCK-8 and plate cloning assays, and migration and invasion abilities were examined with Transwell assays. The pathway regulated by DCLK1 in lung adenocarcinoma was analyzed on the basis of the TCGA lung adenocarcinoma cohort with pathway enrichment analysis and verified through Western blot analysis. Results DCLK1 overexpression in A549 cells promoted cell proliferation, migration, and invasion. The inhibition of the FAK/PI3K/AKT/mTOR signaling pathway impaired the DCLK1-mediated malignant behavior of A549 cells. Conclusion DCLK1 promotes the malignant behavior of A549 cells through the activation of the FAK/PI3K/AKT/mTOR signaling pathway.

Effects and Mechanism of LASP1 on Proliferation, Migration, and Apoptosis of Human Colorectal Cancer LOVO Cells / 肿瘤防治研究

Objective To explore the effects and mechanism of LASP1 gene expression on the proliferation, migration, and invasion of human colorectal cancer (LOVO) cells. Methods LASP1 overexpression plasmids and LASP1 interference plasmids were constructed and transfected to LOVO cells. qRT-PCR was used to detect LASP1 mRNA expression and validate the transfection. MTT method and Tunel staining were used to detect cell proliferation and apoptosis, respectively, and scratch test and Transwell test were employed to determine the migration and invasion abilities of cells. Western blot was applied to analyze the expression of LASP1, p-FAK/FAK, and p-AKT/AKT protein in cells. Results The plasmids were successfully transfected. LASP1 overexpression increased the proliferation, migration, and invasion of LOVO cells, decreased the apoptosis, and increased LASP1, p-FAK/FAK, p-AKT/AKT protein expression (P < 0.01). LASP1 knockdown reduced the proliferation, migration, and invasion of LOVO cells, increased the apoptosis, and decreased LASP1, p-FAK/FAK, and p-AKT/AKT protein expression (P < 0.01). Conclusion LASP1 positively regulates the FAK/AKT signaling pathway to promote the proliferation, migration, and invasion of LOVO cells.

High frequency of alternative splicing variants of the oncogene Focal Adhesion Kinase in neuroendocrine tumors of the pancreas and breast / 医学前沿

The characteristic genetic abnormality of neuroendocrine neoplasms (NENs), a heterogeneous group of tumors found in various organs, remains to be identified. Here, based on the analysis of the splicing variants of an oncogene Focal Adhesion Kinase (FAK) in The Cancer Genome Atlas datasets that contain 9193 patients of 33 cancer subtypes, we found that Box 6/Box 7-containing FAK variants (FAK6/7) were observed in 7 (87.5%) of 8 pancreatic neuroendocrine carcinomas and 20 (11.76%) of 170 pancreatic ductal adenocarcinomas (PDACs). We tested FAK variants in 157 tumor samples collected from Chinese patients with pancreatic tumors, and found that FAK6/7 was positive in 34 (75.6%) of 45 pancreatic NENs, 19 (47.5%) of 40 pancreatic solid pseudopapillary neoplasms, and 2 (2.9%) of 69 PDACs. We further tested FAK splicing variants in breast neuroendocrine carcinoma (BrNECs), and found that FAK6/7 was positive in 14 (93.3%) of 15 BrNECs but 0 in 23 non-NEC breast cancers. We explored the underlying mechanisms and found that a splicing factor serine/arginine repetitive matrix protein 4 (SRRM4) was overexpressed in FAK6/7-positive pancreatic tumors and breast tumors, which promoted the formation of FAK6/7 in cells. These results suggested that FAK6/7 could be a biomarker of NENs and represent a potential therapeutic target for these orphan diseases.

The mechanism of FAK/Twist1 signal pathway in the closure of cranial suture / 安徽医科大学学报

Objective@#To investigate the function of FAK / Twist1 signaling pathway during craniosynostosis closure.@*Methods@#Ten days old rats were divided into a control group (n = 50) and a rotation group (n = 50) . Both of them were made a approximately 0. 5 cm circle bone window at the midpoint of the lambdoid suture of the rat.The bone flaps were left free without damaging the dura mater. The bone flaps in the control group were repositioned in situ , and the bone flaps in the rotation group were rotated 180 ° and repositioned 3 weeks later. Then the experiments were performance as followed :open field test , measurement of body weight , head circumference , bone flap area , and thickness of bone flap in the two groups , observation of cranial suture closure by microscopy and HE staining , FAK / Twist1 expression determined by Western blot , real⁃time PCR , and immunohistochemical staining in the bone flap and dure , respectively. @*Results@#The cranial sutures was completely closed in the rotation groupand that was open in the control group through detecting by microscopic examination and HE staining. The thickness of the bone flap in the derotation group was greater than that in the control group , with statistical significance (P < 0. 01) . There were no significant differences between two groups in head circumference , weight , bone flap area , and operative area. The results of behavioral test showed that after the closure of cranial suture , the acsion of FAK was significantly increased in the calvaria and dura as well as Twsit1 was significantly decreased in the dura in rotating group measuring by Western blot , real⁃time PCR , and immunohistochemical staining (P < 0. 05) .@*Conclusion@#FAK/Twist1 may play an important role in craniosynostosis after rotation.

Study on the mechanism of FAK⁃PI3K/AKT signal pathway in the growth and development of neonatal rat cranial bone / 安徽医科大学学报

Objective@#To explore the growth characteristics of rat calvaria by detecting the calvaria of SD rats in different periods.@*Methods@#The calvaria of SD rats at 1 , 4 , 7 , 10 , and 12 weeks from the same littermate were selected (3 rats per week) . Real⁃time PCR and Western blot techniques were used to detect the expression of focal adhesion kinase ( FAK) Ⅳ phosphatidylinositol 3 kinase/protein kinase B ( PI3K/AKT ) signal pathway in the calvaria , and the role of FAK⁃PI3K/AKT in the growth and development of the calvaria was analyzed by correlation.@*Results@#The increase of brain volume and the thickness of calvaria increased synchronously , the expression of FAK was positively correlated with the changes of meridians , and the expression of FAK was positively correlated with the expression of PI3K/AKT.@*Conclusion@#The expression of FAK is related to the growth and development of rat skull. FAK plays a role in calvaria by activating PI3K/AKT signal pathway. FAK may be used as a marker of rapid skull growth and development , which provides a basic theoretical basis for the timing of clinical skull defect repair and treatment.

Focal adhesion kinase inhibition decreases cell viability and induces apoptosis of JAK2 V617F positive cells

Abstract Focal Adhesion Kinase (FAK) protein participates in proliferation, migration, cell survival, and apoptosis process. It has been described as overexpressed in several neoplasms being a promising target for therapy. BCR-ABL negative chronic Myeloproliferative Neoplasms (MPN) are clonal disorders characterized by the excess of proliferation and apoptosis resistance. The identification of the acquired JAK2 V617F mutation in MPN patients allowed a better understanding of pathogenesis. However, there is still no pharmacological treatment that leads all patients to molecular remission, justifying new studies. The present study aimed to evaluate FAK involvement in the viability and apoptosis of HEL and SET-2 cells, both JAK2 V617F positive cell lines. The FAK inhibitor PF 562,271 was used. Cell viability was determined using MTT assay and apoptosis verified by cleaved PARP, cleaved Caspase 3 and Annexin-V/PI staining detection. FAK inhibition significantly reduced HEL and SET-2 cells viability and induced apoptosis. Considering the role of JAK/STAT pathway in MPN, further investigation of FAK participation in the MPN cells proliferation and apoptosis resistance, as well as possible crosstalk between JAK and FAK and downstream pathways may contribute to the knowledge of MPN pathophysiology, the discovery of new molecular targets, and JAK inhibitors resistance mechanisms.

The Mechanism of BIPN Intervention by Strychnine Based on lncRNA ZFAS1(XIST)/FAK Signaling Pathway / 浙江中医药大学学报

[Objective]To assess the effectiveness of prepared strychnine in the treatment of bortezomib-induced peripheral neuropathy(BIPN)and explore the mechanism of intervention of BIPN based on long noncoding RNA(lncRNA)X inactivated specific transcript(XIST)/ZNFX1 antisense RNA 1(ZFAS1).[Methods]Twenty patients diagnosed as multiple myeloma who received bortezomib(BTZ)and developed BIPN and received strgchnine treatment were collected by prospective non-randomized controlled study method.The traditional Chinese medicine(TCM)symptom score,neurotoxicity score,peripheral neuropathy(PN)grade,and partial peripheral nerve conduction velocity were compared with patients who did not receive strychnine treatment.Using self-control,peripheral blood samples were collected from patients in the treatment group,and enzyme-linked immunosorbent assay(ELISA)was used to detect the expression of inflammation-related factors.DRG 50B11 cells were cultured and screened by cell counting kit-8(CCK-8)for the optimal acting concentration and time of strychnine and the optimal acting time of BTZ,and the cases were randomly divided into normal control group,BTZ group,and strychnine+BTZ group.Real-time quantitative polymerase chain reaction(Real-time qPCR)was used to detect the expression levels of inflammation-related factors and total RNA related indexes,and it analyzed the differences and correlations.[Results]The clinical study showed that compared with control group,PN,TCM syndrome scores and neurotoxicity score were decreased after treatment,while peripheral nerve conduction velocity was increased(P＜0.05),and there were no significant adverse effects.The experimental results showed that compared with those before treatment,the expression of interleukin-17(IL-17),tumor necrosis factor-α(TNF-α),IL-1β,IL-6,nerve growth factor(NGF)and brain-derived neurotrophic factor(BDNF)were reduced(P＜0.05),and there was a significant negative correlation with time(P＜0.01).Compared with BTZ group,the expression levels of IL-17,TNF-α,IL-1β,IL-6,NGF,BDNF,the lncRNA XIST,fibronectin 1(FN1)and phospho-focal adhesion kinase(p-FAK)were decreased in strychnine+BTZ group(P＜0.05),while the expressions of miR-96-5P and miR-1271-5P increased(P＜0.05),without significant difference in the expression of lncRNA ZFAS1(P＞0.05).lncRNA XIST expression levels were significantly positively correlated with the expressions of IL-17,TNF-α,IL-1β,IL-6,NGF,BDNF,FN1 and p-FAK(P＜0.01),but no moderate negative correlated with miR-96-5P(P＜0.05),or very weakly correlated or no correlated with miR-1271-5P(P＞0.05).[Conclusion]Prepared strychnine capsule can alleviate BIPN to a certain extent and is relatively safe,and its mechanism may be related to the regulation of lncRNA XIST for promoting the expression of miR-96-5P/FN1 and inhibit p-FAK-mediated neuroinflammation.

Effect of Banxia Xiexintang-containing Intestinal Absorption Solution on Migration and Invasion of PMN-MDSCs in Gastric Cancer Microenvironment / 中国实验方剂学杂志

ObjectiveTo observe the effect of Banxia Xiexintang containing intestinal absorption solution （BXCIAS） on migration and invasion of polymorphonuclear myeloid-derived suppressor cells （PMN-MDSCs） in gastric cancer microenvironment. MethodThe complex solution （containing 0.63 g·mL-1 crude drug） was prepared. Gastric cancer cells were subjected to non-contact co-culture with PMN-MDSCs in Transwell chamber to create gastric cancer microenvironment. Cell counting kit-8 （CCK-8） assay was used to screen the optimal intervention concentration and time of BXCIAS on PMN-MDSCs for subsequent experiment. The blank group， model group， FAK inhibitor group， and BXCIAS groups （26%， 18%， and 10%） were designed. Scratch assay and Transwell assay were employed to detect the migration and invasion ability of PMN-MDSCs， and enzyme-linked immunosorbent assay （ELISA） to measure the expression of vascular endothelial cell growth factor （VEGF） and matrix metalloproteinase-9 （MMP-9） in tumor microenvironment. The expression levels of PMN-MDSCs pathway-related proteins FAK， phosphorylated （p）-FAK， protein tyrosine kinase （Src）， and p-Src were detected by Western blot. ResultThe inhibition rates of PMN-MDSCs by 5%， 50%， 75%， and 100% BXCIAS at 48 h were higher than those at 24 h （P<0.05， P<0.01）. The inhibition rate of PMN-MDSCs by 50% BXCIAS at 72 h was lower than that at 48 h （P<0.01）， and the inhibition rates by 5% and 100% BXCIAS at 72 h were higher than those at 48 h （P<0.05， P<0.01）. There was no significant difference in the inhibition rate by other concentration levels at 48 h. The half-maximal inhibitory concentration （IC50） at 48 h was 18.09%， indicating that 18% BXCIAS and 48 h were the optimal concentration and time， respectively. The migration distance of PMN-MDSCs was large （P<0.01）， and the number of migrating and invading cells increased （P<0.01） in the mode group compared with those in the blank group. Compared with model group， FAK inhibitor and BXCIAS at different concentration decreased the migration distance of PMN-MDSCs （P<0.01）， and the number of migrating and invading cells （P<0.01）， especially the 26% BXCIAS （P<0.01）. The expression of PMN-MDSCs pathway-related proteins FAK， p-FAK， Src and p-Src （P<0.01） and the expression of VEGF and MMP-9 （P<0.01） were higher in the model group than in the blank group. Compared with model group， FAK inhibitor and BXCIAS （26%， 18%， 10%） decreased the expression of FAK， p-FAK， and Src （P<0.01）， and FAK inhibitor and 18% BXCIAS reduced the expression of p-Src （P<0.01）， and the expression of VEGF and MMP-9 （P<0.01）. ConclusionBXCIAS can inhibit the migration and invasion of PMN-MDSCs by down-regulating the expression of FAK， p-FAK， Src， and p-Src proteins in the FAK signaling pathway of PMN-MDSCs in gastric cancer microenvironment.

Baicalein Mediated FAK Protein Regulates PI3K/Akt Signaling Pathway to Inhibit Proliferation and Migration of Gastric Cancer HGC-27 Cells / 中国实验方剂学杂志

ObjectiveTo study the possible molecular mechanism of baicalein （BAI）-mediated focal adhesion kinase （FAK） in the regulation of phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway to inhibit the proliferation and migration of gastric cancer HGC-27 cells. MethodThe gastric epithelial GES-1 cells and gastric cancer HGC-27 cells were respectively treated with BAI （0， 5， 15， 25， and 50 μmol·L-1） for 48 h， and then methyl thiazolyl tetrazolium （MTT） assay was adopted to detect effect of BAI on cell proliferation. Western blot （WB） was employed to detect the expression of FAK and the proteins related to epithelial-mesenchymal transition （EMT） and PI3K signaling pathway after intervention with different concentrations of BAI. The HGC-27 cells stably overexpressing FAK were constructed with lentivirus-mediated transfection technique， and the transfection of FAK was detected through WB and green fluorescent protein （GFP）. The cells were divided into empty vector （NC） group， BAI group， FAK overexpression group， and BAI-treated FAK overexpression group， and cell proliferation activity was detected by MTT assay. The colony formation and cell migration were observed via colony formation assay and Transwell migration assay， respectively. The expression of proteins involved in EMT and PI3K signaling pathways were detected by Western blot. ResultCompared with the NC group， BAI （15， 25 and 50 μmol·L-1） inhibited the proliferation of HGC-27 cells in a dose-dependent manner （P<0.05， P<0.01） while did not affect that of GES-1 cells. BAI （5， 15 and 25 μmol·L-1） down-regulated the expression level of p-FAK （P<0.05， P<0.01）. Compared with NC group， FAK overexpression group showed up-regulated expression level of FAK in HGC-27 cells. The HGC-27 cells in both NC group and FAK overexpression group had green fluorescence. Compared with NC group， BAI inhibited the growth， colony formation， and migration， while FAK overexpression promoted those of HGC-27 cells. The treatment of FAK overexpression group with BAI inhibited the enhancement of cell proliferation and migration （P<0.05）. WB showed that compared with NC group， BAI （15， 25 μmol·L-1） significantly up-regulated the expression of E-cadherin protein and down-regulated that of Vimentin， Snail， p-PI3K， and p-Akt protein in HGC-27 cells （P<0.05， P<0.01）. Compared with NC group， FAK overexpression group showed down-regulated expression of E-cadherin， up-regulated expression of p-FAK， Vimentin， and Snail， and increased ratios of p-FAK/FAK， p-PI3K/PI3K and p-Akt/Akt （P<0.05）. This phenomenon would be reversed after BAI treatment. ConclusionBAI can affect the proliferation and migration of gastric cancer HGC-27 cells by mediating FAK to regulate PI3K/Akt signaling pathway.

CTHRC1 promotes the migration and invasion of bladder cancer 5637 cells through activating the FAK-ERK1/2 signaling pathway / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：探讨胶原三螺旋重复蛋白1（CTHRC1）在膀胱癌组织和细胞中的表达及其对膀胱癌5637细胞迁移和侵袭的影响及其机制。方法：利用TCGA和Arrayexpress数据库中膀胱癌基因表达数据，分析CTHRC1转录和翻译水平。收集2014年9月至2020年12月重庆医科大学附属第一医院手术切除的144例膀胱癌组织和25例全膀胱切除的癌旁组织标本，以及人膀胱癌细胞RT4、5637、T24、UMUC-3、TCCSUP和输尿管上皮永生化细胞SV-HUC-1。采用免疫组织化学染色法、qPCR法和WB法检测膀胱癌组织和细胞中CTHRC1的表达水平，通过Kaplan-Meier曲线分析CTHRC1表达对总生存期（OS）的影响。运用RNAi技术，敲降5637细胞CTHRC1表达后，通过细胞划痕实验和Transwell实验检测CTHRC1表达下调对5637细胞迁移和侵袭的影响。利用基因集富集分析（GSEA）预测CTHRC1相关的潜在信号通路，WB法检测敲降CTHRC1表达对FAK-ERK1/2通路相关蛋白表达的影响。结果：CTHRC1的转录和翻译水平在肌层浸润性膀胱癌（MIBC）组织和细胞中表达显著上调（均P<0.05），CTHRC1高表达组患者5年OS较低表达患者缩短（P<0.05）。干扰CTHRC1表达后，膀胱癌5637细胞迁移及侵袭能力均显著降低（均P<0.01）。GSEA预测显示，CTHRC1高表达组主要富集在黏着斑激酶（FAK）、肌动蛋白细胞骨架调节、FAK和ERK1/2信号通路。WB法实验结果表明，重组CTHRC1蛋白促进膀胱癌5637细胞FAK-ERK1/2信号通路活化（P<0.05或P<0.01）。结论：CTHRC1在MIBC中表达上调，且与膀胱癌患者不良预后密切相关；CTHRC1促进膀胱癌细胞迁移和侵袭，该过程可能与FAK-ERK1/2信号通路的激活有关。

TMEM16A inhibits angiotensin II-induced basilar artery smooth muscle cell migration in a WNK1-dependent manner

Vascular smooth muscle cell (VSMC) migration plays a critical role in the pathogenesis of many cardiovascular diseases. We recently showed that TMEM16A is involved in hypertension-induced cerebrovascular remodeling. However, it is unclear whether this effect is related to the regulation of VSMC migration. Here, we investigated whether and how TMEM16A contributes to migration in basilar artery smooth muscle cells (BASMCs). We observed that AngII increased the migration of cultured BASMCs, which was markedly inhibited by overexpression of TMEM16A. TMEM16A overexpression inhibited AngII-induced RhoA/ROCK2 activation, and myosin light chain phosphatase (MLCP) and myosin light chain (MLC20) phosphorylation. But AngII-induced myosin light chain kinase (MLCK) activation was not affected by TMEM16A. Furthermore, a suppressed activation of integrin

Role of FAK signaling in chagasic cardiac hypertrophy

Abstract Cardiac hypertrophy and dysfunction are a significant complication of chronic Chagas disease, with heart failure, stroke, and sudden death related to disease progression. Thus, understanding the signaling pathways involved in the chagasic cardiac hypertrophy may provide potential targets for pharmacological therapy. Herein, we investigated the implication of focal adhesion kinase (FAK) signaling pathway in triggering hypertrophic phenotype during acute and chronic T. cruzi infection. C57BL/6 mice infected with T. cruzi (Brazil strain) were evaluated for electrocardiographic (ECG) changes, plasma levels of endothelin-1 (ET-1) and activation of signaling pathways involved in cardiac hypertrophy, including FAK and ERK1/2, as well as expression of hypertrophy marker and components of the extracellular matrix in the different stages of T. cruzi infection (60-210 dpi). Heart dysfunction, evidenced by prolonged PR interval and decrease in heart rates in ECG tracing, was associated with high plasma ET-1 level, extracellular matrix remodeling and FAK signaling activation. Upregulation of both FAK tyrosine 397 (FAK-Y397) and serine 910 (FAK-S910) residues phosphorylation as well as ERK1/2 activation, lead to an enhancement of atrial natriuretic peptide gene expression in chronic infection. Our findings highlight FAK-ERK1/2 signaling as a regulator of cardiac hypertrophy in Trypanosoma cruzi infection. Both mechanical stress, induced by cardiac extracellular matrix (ECM) augment and cardiac overload, and ET-1 stimuli orchestrated FAK signaling activation with subsequent activation of the fetal cardiac gene program in the chronic phase of infection, highlighting FAK as an attractive target for Chagas disease therapy.

Anastrozole promotes implantation by altering the expression of paxillin and fak in rat luminal uterine epithelium / El anastrozol promueve la implantación alterando la expresión de paxilina y fak en el epitelio uterino luminal de rata

An alternative hyper-ovulator inducer to replace clomiphene citrate (CC) is needed as it is unsuitable for women with polycystic ovarian syndrome and is associated with low pregnancy rates. Anastrozole is an effective hyper-ovulator inducer, but has not been well researched. In order to determine the effectiveness of anastrozole as a hyper-ovulator inducer and to an extent compare it with CC in similar situations, this study ascertained the effects of these drugs on the expression of the focal adhesion proteins, paxillin and FAK, which are uterine receptivity markers in the surface luminal uterine epithelial cells of day 1 and day 6 pregnant Wistar rats. The results show that paxillin is localized in focal adhesions at the base of the uterine epithelial cells at day 1 of pregnancy whereas at day 6, paxillin disassembles from the basal focal adhesions and localizes and increases its expression apically. FAK is faintly expressed at the basal aspect of the uterine epithelial cells while moderately expressed at the cell-to-cell contact at day 1 in all groups from where it disassembles and relocates apically and becomes more intensely expressed at day 6 of pregnancy in untreated and anastrozole treated rats. Although paxillin is localized apically at day 6, its expression is significantly down-regulated with CC treatment suggesting its interference with the implantation process. These findings seem to suggest that anastrozole could favor implantation.

Para reemplazar el citrato de clomifeno (CC) es necesario un inductor de hiperovulación alternativo, ya que no es adecuado para mujeres con síndrome de ovario poliquístico y está asociado con tasas bajas de embarazo. El anastrozol es un inductor eficaz del hiper-ovulador, pero no se ha investigado adecuadamente. Con el fin de determinar la efectividad del anastrozol como inductor del hiper-ovulador y, en cierta medida, compararlo con CC en situaciones similares, este estudio determinó los efectos de estos fármacos en la expresión de las proteínas de adhesión focal, paxillin y FAK, uterinas marcadores de receptividad en la superficie luminal de células uterinas epiteliales, del día 1 y día 6 en ratas Wistar preñadas. Los resultados muestran que la paxilina se localiza en adherencias focales en la base de las células epiteliales uterinas en el día 1 del embarazo, mientras que en el día 6, la paxilina se desmonta de las adherencias focales basales y localiza y aumenta su expresión apicalmente. FAK se expresa débilmente en el aspecto basal de las células epiteliales uterinas, mientras que se expresa moderadamente en el contacto de célula a célula en el día 1 en todos los grupos, donde se separa y se reubica apicalmente y se expresa con mayor intensidad el día 6 de la preñez, en pacientes no tratados y tratados. ratas tratadas con anastrozol. Aunque la paxillina se localiza apicalmente en el día 6, su expresión está significativamente disminuida con el tratamiento con CC, lo que sugiere su interferencia con el proceso de implantación. Estos hallazgos sugieren que el anastrozol podría favorecer el proceso de implantación.

Afzelin inhibits migration of MDA-MB-231 cells by suppressing FAK expression and Rac1 activation

Triple-negative breast cancer (TNBC) has the worst prognosis and the highest rate of metastasis among other types ofbreast cancer. These characteristics are supported by the dysregulation of focal adhesion kinase (FAK) and Rac1 whichare the key players of mesenchymal cell migration on TNBC. Afzelin is a secondary metabolite that is contained ina variety of plants. This study explored the anti-migration effect of afzelin and its interaction with FAK and Rac1 onthe highly invasive TNBC cell line, MDA-MB-231. Cell viability was assessed by 3-(4,5-dimethyl 2-thiazolyl)-2,5-diphenyltetrazolium bromide assay, and cell migration was evaluated using in vitro scratch assay. Rac1 activation wasanalyzed using the colorimetric assay, while vinculin and actin filaments were stained through immunofluorescence. Thequantity of total FAK and phosphorylated FAK tyr397 was detected by Western blotting. Afzelin decreased cell viabilityand inhibited two-dimensional cell migration in a dose-dependent manner. Under confocal laser scanning microscopy,vinculin localization at the cell edge demonstrated a reduction of focal adhesion formation by afzelin. Further explorationshowed that afzelin decreased FAK expression but did not affect FAK phosphorylation at tyr397. In addition, afzelindecreased Rac1-GTPase activation, which is a downstream effector of FAK. Taken together, these results suggest thatafzelin suppresses TNBC cell migration, through inhibition of FAK expression and Rac1-GTPase activation.

P38/akt pathway regulates the oriented differentiation of bone marrow mesenchymal stem cells in nanofiber annulus fibrosus scaffolds with different spatial structures / 中国组织工程研究

BACKGROUND: Previous studies have shown that a variety of materials can be used for the construction of tissue engineering scaffolds. The topological structure of the scaffold surface has a regulatory effect on the biological behaviors such as stem cell proliferation and differentiation, but the specific mechanism is still unclear. OBJECTIVE: To investigate the role of P38 and Akt pathways in the oriented differentiation of bone marrow mesenchymal stem cells in nanofiber scaffolds. METHODS: Three kinds of nanofiber scaffolds (AFS, AYS, 3-DPS) with different structures were constructed. Rat bone marrow mesenchymal stem cells were inoculated on the surface of three kinds of nanofiber scaffolds. After osteogenic induction, cell morphology, adhesion and proliferation were detected. mRNA expression levels of key phenotype molecules (COLIα1, COLIIα1, Aggrecan, Sox-9) were measured using qRT-PCR. Intracellular P38, AKT, ERK1/2 and JNK expression was detected by western blot assay. RESULTS AND CONCLUSION: After 4 and 8 hours of culture, cell adhesion rate of the 13-DPS scaffold group was higher than that of the AFS and AYS scaffold groups (P<0.05). After 7 days of culture, cells of the 13-DPS scaffold group proliferated faster than those of AFS and AYS scaffold groups (P<0.05). Bone marrow mesenchymal stem cells adhered firmly and grew well on three kinds of scaffolds. Fibroblast-like growth was observed on the AFS and AYS scaffolds and chondrocyte-like growth was observed on the 3-DPS scaffold. After 3 weeks of cartilage induction, mRNA expression of COLIIα1, Aggrecan and Sox-9 was higher, and the mRNA expression of COLIα1 was lower, in the 3-DPS scaffold group compared with the other two groups (both P<0.05). After 3 weeks of cartilage induction, relative expression level of p-AKT and p-P38 in the 3-DPS scaffold group was significantly higher than that in the other two groups (both P<0.05). There were no significant differences in AKT total protein and ERK1/2, JNK, P38, p-ERK1/2, p-JNK and p-P38 protein expression levels among three groups. These findings suggest that nanofiber annulus fibrosus scaffolds with different spatial structures can induce the oriented differentiation of bone marrow mesenchymal stem cells through the P38 and AKT pathway, which were the downstream of the Integrin-FAK signaling pathway.

Extracellular signal regulated kinase 5 promotes cell migration, invasion and lung metastasis in a FAK-dependent manner

This study was designed to evaluate ERK5 expression in lung cancer and malignant melanoma progression and to ascertain the involvement of ERK5 signaling in lung cancer and melanoma. We show that ERK5 expression is abundant in human lung cancer samples, and elevated ERK5 expression in lung cancer was linked to the acquisition of increased metastatic and invasive potential. Importantly, we observed a significant correlation between ERK5 activity and FAK expression and its phosphorylation at the Ser

PEP06 polypeptide 30 is a novel cluster-dissociating agent inhibiting v integrin/FAK/Src signaling in oral squamous cell carcinoma cells

Collectively migrating tumor cells have been recently implicated in enhanced metastasis of epithelial malignancies. In oral squamous cell carcinoma (OSCC), v integrin is a crucial mediator of multicellular clustering and collective movement ; however, its contribution to metastatic spread remains to be addressed. According to the emerging therapeutic concept, dissociation of tumor clusters into single cells could significantly suppress metastasis-seeding ability of carcinomas. This study aimed to investigate the anti-OSCC potential of novel endostatin-derived polypeptide PEP06 as a cluster-dissociating therapeutic agent . Firstly, we found marked enrichment of v integrin in collectively invading multicellular clusters in human OSCCs. Our study revealed that metastatic progression of OSCC was associated with augmented immunostaining of v integrin in cancerous lesions. Following PEP06 treatment, cell clustering on fibronectin, migration, multicellular aggregation, anchorage-independent survival and colony formation of OSCC were significantly inhibited. Moreover, PEP06 suppressed v integrin/FAK/Src signaling in OSCC cells. PEP06-induced loss of active Src and E-cadherin from cell-cell contacts contributed to diminished collective migration of OSCC . Overall, these results suggest that PEP06 polypeptide 30 inhibiting v integrin/FAK/Src signaling and disrupting E-cadherin-based intercellular junctions possesses anti-metastatic potential in OSCC by acting as a cluster-dissociating therapeutic agent.