Dihydroartemisinin inhibits HNSCC invasion and migration by controlling miR-195-5p expression.

Objectives: Dihydroartemisinin (DHA), an artemisinin derivative extracted from the traditional Chinese medicinal herb Artemisia annua, has the potential to suppress head and neck squamous cell carcinoma (HNSCC) progression. However, the mechanisms underlying these effects remain unclear. Therefore, we aimed to examine the mechanisms underlying the effects of DHA on tumor invasion and migration. Methods: Human HNSCC cell lines CAL-27 and FaDu were exposed to varying DHA concentrations (0, 5, 20, and 80 µM) for 24 h. Cell proliferation, invasion, and migration were assessed using CCK8, transwell, and wound-healing assays, respectively. Quantitative real-time PCR, western blotting, and immunofluorescence were used to assess the expression levels of the target genes and proteins. Results: DHA suppressed the invasion and migration of CAL-27 and FaDu cells. Additionally, miR-195-5p suppressed the invasion and migration of HNSCC cells. This study revealed significant differences in the expression of miR-195-5p and TENM2 between clinical samples and multiple public databases. DHA treatment and miR-195-5p overexpression significantly reduced TENM2 expression in HNSCC cells, which suggested that miR-195-5p overexpression enhanced the inhibitory effect of DHA on TENM2. Conclusions: This study provides the first evidence that DHA inhibits cell invasion and migration by regulating the miR-195-5p/TENM2 axis in HNSCC cells, suggesting it as a potentially effective treatment strategy for HNSCC.

Identification of a Novel miR-195-5p/PNN Axis in Colorectal Cancer.

Pinin (PNN) is a desmosome-associated protein that reinforces the organization of keratin intermediate filaments and stabilizes the anchoring of the cytoskeleton network to the lateral surface of the plasma membrane. The aberrant expression of PNN affects the strength of cell adhesion as well as modifies the intracellular signal transduction pathways leading to the onset of CRC. In our previous studies, we characterized the role of miR-195-5p in the regulation of desmosome junctions and in CRC progression. Here, with the aim of investigating additional mechanisms related to the desmosome complex, we identified PNN as a miR-195-5p putative target. Using a public data repository, we found that PNN was a negative prognostic factor and was overexpressed in colon cancer tissues from stage 1 of the disease. Then, we assessed PNN expression in CRC tissue specimens, confirming the overexpression of PNN in tumor sections. The increase in intracellular levels of miR-195-5p revealed a significant decrease in PNN at the mRNA and protein levels. As a consequence of PNN regulation by miR-195-5p, the expression of KRT8 and KRT19, closely connected to PNN, was affected. Finally, we investigated the in vivo effect of miR-195-5p on PNN expression in the colon of AOM/DSS-treated mice. In conclusion, we have revealed a new mechanism driven by miR-195-5p in the regulation of desmosome components, suggesting a potential pharmacological target for CRC therapy.

Role of miR-128-3p and miR-195-5p as biomarkers of coronary artery disease in Indians: a pilot study.

Coronary artery disease (CAD) imposes a significant economic burden in developing countries like India. Timely diagnosis and treatment should be prioritized to mitigate the disease. Current diagnostic tools being invasive and less specific raise the need to develop less invasive and more reliable molecular biomarkers. MicroRNAs (miRNAs) are an emerging class of molecules that can serve as a potential source of non-invasive biomarkers for CAD. The objective of this study was to determine the potential of circulatory miRNAs as diagnostic biomarkers in CAD. In this study, we have reported two microRNAs, miR-128-3p and miR-195-5p in the serum of CAD patients in Indian Population. A total of 124 subjects were recruited which included 89 angiographically proven CAD patients and 35 control subjects. Our results show a significant decrease in the levels of miR-128-3p in CAD patients while there were no significant changes in the levels of miR-195-5p. Further bioinformatics analysis revealed the potential role of miR-128-3p in cholesterol homeostasis. Altered homeostasis due to cholesterol accumulation in macrophages is the driving force behind formation of foam cells which in turn accelerates the progression of CAD. Here, we have shown that miR-128-3p increases cholesterol levels in macrophages by decreasing cholesterol efflux in-vitro.

Mir-195-5p targets Smad7 regulation of the Wnt/ß-catenin pathway to promote osteogenic differentiation of vascular smooth muscle cells.

In this study, we sought to investigate the mechanisms of action of miR-195-5p in the osteogenic differentiation of vascular smooth muscle cells (VSMCs), and thereby provide novel insights and a reference for the targeted therapy of arterial media calcification. VSMC differentiation was induced using sodium ß-glycerophosphate, and we investigated the effects of transfecting cells with miR-195-5p mimics, vectors overexpressing Smad7, and the Wnt/ß-catenin pathway inhibitor (KYA1797K) on VSMC differentiation by determining cell viability and apoptosis, and the mRNA and protein expression of factors associated with osteogenic differentiation and the Wnt/ß-catenin pathway. The results revealed that miR-195-5p mimics enhanced the osteogenic differentiation of VSMCs induced by ß-glycerophosphate, whereas the overexpression of Smad7 reversed this phenomenon. In addition, KYA1797K was found to promote the effects of Smad7 overexpression. In conclusion, by targeting, Smad7, miR-195-5p promotes the Wnt/ß-catenin pathway. and thus the osteogenic differentiation of VSMCs. These findings will provide a reference for elucidating the mechanisms whereby miR-195-5p regulates osteogenic differentiation.

Exosome-transported of circ_0081069 induces SPIN1 production by binding to miR-195-5p to inhibit radiosensitivity in esophageal squamous cell carcinoma.

Circ_0081069 plays a key role in tumor growth; however, its effect on radiosensitivity in esophageal squamous cell carcinoma (ESCC) remains unknown. The study is performed to reveal the association of circ_0081069 expression and radiosensitivity in ESCC and the underlying mechanism. Circ_0081069, miR-195-5p, and spindlin 1 (SPIN1) RNA expression were detected by quantitative real-time polymerase chain reaction. Protein expression was checked by Western blot analysis or immunohistochemistry assay. Cell viability, proliferation, cell apoptosis, migration, and invasion were investigated by cell counting kit-8, 5-Ethynyl-29-deoxyuridine, flow cytometry analysis, scratch test, and transwell assays, respectively. The sensitivity of ESCC cells to radiation was investigated by cell colony formation assay. The interactions among circ_0081069, miR-195-5p, and SPIN1 were identified by dual-luciferase reporter assay and RNA Immunoprecipitation assay. Xenograft mouse model assay was performed to determine the effect of circ_0007841 on radiosensitivity in vivo. Circ_0081069 and SPIN1 expression were upregulated, whereas miR-195-5p was downregulated in ESCC tissues, ESCC cells, and radiation-stimulated ESCC cells. Circ_0081069 silencing inhibited ESCC cell proliferation, invasion, and migration but improved cell apoptosis. In addition, circ_0081069 knockdown enhanced ESCC cell radiosensitivity in vitro and in vivo. Circ_0081069 bound to miR-195-5p and regulated radiosensitivity by binding to miR-195-5p in ESCC cells. Moreover, SPIN1, a target of miR-195-5p, rescued miR-195-5p-mediated effects in ESCC cells. Circ_0081069 was secreted from ESCC cells by being packaged into exosomes. Further, circ_0081069-Exo inhibited radiosensitivity in ESCC cells. Exosome-mediated transfer of circ_0081069 induced SPIN1 production by binding to miR-195-5p, further inhibiting radiosensitivity in ESCC.

Hsa-miR-194-5p and hsa-miR-195-5p are down-regulated expressed in high dysplasia HPV-positive Pap smear samples compared to normal cytology HPV-positive Pap smear samples.

BACKGROUND: The human papillomavirus (HPV) infection may affect the miRNA expression pattern during cervical cancer (CC) development. To demonstrate the association between high-risk HPVs and the development of cervix dysplasia, we examined the expression patterns of hsa-miR-194-5p and hsa-miR-195-5p in Pap smear samples from southeast Iranian women. We compared samples that were HPV-positive but showed no abnormality in the cytological examination to samples that were HPV-positive and had severe dysplasia. METHODS: Pap smear samples were obtained from 60 HPV-positive (HPV-16/18) patients with histologically confirmed severe dysplasia (cervical intra-epithelial neoplasia (CIN 3) or carcinoma in situ) and the normal cytology group. The expression of hsa-miR-194-5p and hsa-miR-195-5p was analyzed by real-time quantitative PCR, using specific stem-loop primers and U6 snRNA as the internal reference gene. Clinicopathological features were associated with miRNA expression levels. Furthermore, functional enrichment analysis was conducted using in silico tools. The Kaplan-Meier survival method was also obtained to discriminate survival-significant candidate miRNAs in CC, and receiver operating characteristic (ROC) curves were constructed to assess the diagnostic value. RESULTS: Compared to HPV-positive cytologically normal Pap smear samples, hsa-miR-194-5p and hsa-miR-195-5p relative expression decreased significantly in HPV-positive patients with a severe dysplasia Pap smear. Kaplan-Meier analysis indicated a significant association between the miR-194 decrease and poor CC survival. In essence, ROC curve analysis showed that miR-194-5p and miR-195-5p could serve as valuable markers for the development of cervix dysplasia in individuals who are positive for high-risk HPVs. CONCLUSIONS: This study revealed that hsa-miR-194-5p and hsa-miR-195-5p may possess tumor suppressor capabilities in the context of cervical dysplasia progression. However, it remains uncertain whether these microRNAs are implicated in the transition of patients with high dysplasia to cervical cancer. We also showed the potential capability of candidate miRNAs as novel diagnostic biomarkers related to cervical dysplasia progression.

MicroRNA-195-5p Attenuates Pregnancy-Induced Hypertension by Inhibiting Oxidative Stress via OTX1/MAPK Signaling Pathway.

Pregnancy-induced hypertension (PIH) is a hypertensive disorder during pregnancy and can induce perinatal death of human infants. MicroRNA (miR)-195-5p was validated to display low expression in severe preeclampsia placentas, but the role of miR-195-5p in pregnancy-induced hypertension (PIH) has not been investigated. The study emphasized on the functions and mechanism of miR-195-5p in PIH. A reduced uterine perfusion pressure (RUPP) rat model was established to mimic PIH in vivo. Adenovirus (Ad)-miR-195-5p agomir and/or Ad-OTX1 were further injected into some model rats. RT-qPCR was conducted to assess the expression of miR-195-5p and orthodenticle homeobox 1 (OTX1) in rat placental tissues, the isolated aortic endothelial cells (AECs), and in serum samples of PIH patients. Western blot analysis was implemented to measure the protein levels of OTX1, VEGFA, and key factors involved in the MAPK signaling pathway. The concentrations of oxidative stress markers (superoxide dismutase, catalase, and lipid hydroperoxide) in AECs and placental tissues of RUPP rats were measured by corresponding kits. The binding relation between miR-195-5p and OTX1 was verified using the dual-luciferase reporter assay. Hematoxylin-eosin staining was conducted to evaluate the pathological features of rat placental tissues. MiR-195-5p was downregulated, while OTX1 was upregulated in rat placental tissues and human serum samples of PIH patients. MiR-195-5p could target OTX1 and inversely regulate OTX1 expression in AECs and rat placental tissues. In addition, miR-195-5p can negatively regulate VEGFA level. Furthermore, miR-195-5p inactivates oxidative stress and the MAPK signaling by downregulating OTX1 in AECs. In vivo experiments revealed that OTX1 overexpression reversed the protective effect of miR-195-5p overexpression on placental damage and oxidative stress. MiR-195-5p alleviates PIH by inhibiting oxidative stress via targeting OTX1 and inactivating MAPK signaling.

The regulatory effect of lncRNA LINC00943 on the progression of hepatocellular carcinoma and its relationship with clinicopathological features.

BACKGROUND: The risk factors for the pathogenesis of HCC are highly variable, and the prognosis of patients is very unsatisfactory. In this study, we investigated the regulatory effect of LINC00943 on HCC progression and its relationship with clinicopathological features. METHODS: LINC00943 level in HCC tissues and cell specimens was verified by RT-qPCR. The pathologic significance of LINC00943 in the prognosis of HCC was analyzed by Kaplan-Meier and Cox regression analyses. The behavioral function of LINC00943 in HCC cells was evaluated via CCK-8 and Transwell assays. The specific targeting relationship between LINC00943 and miR-195-5p was investigated by luciferase activity assay. RESULTS: LINC00943 was highly expressed in HCC tissues and cell specimens. Clinical data analysis showed that elevated LINC00943 indicated poor prognosis in patients with HCC and was related to TNM stage and lymph node metastasis. Cell experiments demonstrated that silencing LINC00943 sponge miR-195-5p suppressed the proliferation, migration and invasion of HCC cells. Mechanistically, miR-195-5p inhibitor remedied the suppressive effect of silencing LINC00943 on the biological functions of HCC cells. CONCLUSION: LINC00943 may be an independent prognostic factor of HCC, which provides new thinking for the prognosis and treatment of HCC patients.

miR-195-5p as Regulator of γ-Catenin and Desmosome Junctions in Colorectal Cancer.

Desmosomes play a key role in the regulation of cell adhesion and signaling. Dysregulation of the desmosome complex is associated with the loss of epithelial cell polarity and disorganized tissue architecture typical of colorectal cancer (CRC). The aim of this study was to investigate and characterize the effect of miR-195-5p on desmosomal junction regulation in CRC. In detail, we proposed to investigate the deregulation of miR-195-5p and JUP, a gene target that encodes a desmosome component in CRC patients. JUP closely interacts with desmosomal cadherins, and downstream, it regulates several intracellular transduction factors. We restored the miR-195-5p levels by transient transfection in colonic epithelial cells to examine the effects of miR-195-5p on JUP mRNA and protein expression. The JUP regulation by miR-195-5p, in turn, determined a modulation of desmosome cadherins (Desmoglein 2 and Desmocollin 2). Furthermore, we focused on whether the miR-195-5p gain of function was also able to modulate the expression of key components of Wnt signaling, such as NLK, LEF1 and Cyclin D1. In conclusion, we have identified a novel mechanism controlled by miR-195-5p in the regulation of adhesive junctions, suggesting its potential clinical relevance for future miRNA-based therapy in CRC.

Effect of lncRNA LINC00324 on cervical cancer progression through down-regulation of miR-195-5p.

BACKGROUND: Long non-coding RNAs (lncRNAs) have been widely used in the exploration of diseases in recent years. This paper introduced the significance of lncRNA LINC00324 (LINC00324) on the progression of cervical cancer and explored the mechanism of action and potential prognosis of LINC00324. METHODS: The cervical cancer tissues and adjacent normal tissues of 120 people were collected as research samples. The expression level of LINC00324 was assessed by RT-qPCR, as was miR-195-5p. Knockdown of LINC00324 on the proliferation ability of cervical cancer cells was determined with the help of cell counting kit-8 (CCK-8), and the number of cell migration and invasion was detected by the Transwell method. Luciferase reporter gene assay was used to analyse the correlation of LINC00324 and miR-195-5p. Kaplan-Meier survival curves and multivariate Cox analysis explained the potential prognostic significance of LINC00324 in cervical cancer. RESULTS: Significantly increased expression of LINC00324 and down-regulated miR-195-5p were negatively correlated in cervical cancer. Knockdown of LINC00324 inhibited the progression of cervical cancer, which was related to its mechanism of targeting and downregulating miR-195-5p. In addition, low expression of LINC00324 may prolong the survival period of patients with cervical cancer. CONCLUSIONS: LINC00324 targets miR-195-5p to regulate the progression of cervical cancer and have potential as a prognostic molecular marker for cervical cancer.

This paper introduced the mechanism and prognostic potential of LINC00324 in cervical cancer. The study found that LINC00324 expression was significantly elevated, while miR-195-5p level was down-regulated in cervical cancer. LINC00324 sponging miR-195-5p regulated the proliferation, migration and invasion of cervical cancer cells, thereby affecting the progression of cervical cancer. LINC00324 may be a prognostic biomarker for cervical cancer, providing a new direction for the treatment of patients.

ATG9A modulated by miR-195-5p can boost the malignant progression of cervical cancer cells.

Cervical cancer (CC) is a major public health problem, and its molecular mechanism requires further investigation. The goal of this study was to determine the role of miR-195-5p and the autophagy-related protein ATG9A in tumour metastasis, epithelial - mesenchymal transition (EMT), apoptosis, and autophagy of CC cells. Using bioinformatics analysis, we predicted ATG9A as a downstream target gene of miR-195-5p, an integral membrane protein required for autophagosome formation and involved in tumorigenesis. Next, western blotting and Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) showed that upregulation of miR-195-5p decreased protein and mRNA expression of ATG9A, and downregulation of miR-195-5p promoted ATG9A protein and mRNA expression. In addition, detection of the dual luciferase reporter gene further indicated ATG9A is a direct downstream target gene of miR-195-5p. Finally, the effects of miR-195-5p and ATG9A on CC cell proliferation, migration, invasion, EMT, autophagy, and apoptosis were evaluated in vitro. Our results showed that upregulation of miR-195-5p not only inhibits proliferation, migration, and the EMT of CC cells, but also induces apoptosis and autophagy. Conversely, downregulation of miR-195-5p increased malignant metastasis and the EMT of CC cells, and inhibited apoptosis as well as autophagy. In addition, miR-195-5p targeted and negatively regulated ATG9A, and rescue experiments suggested that overexpression of ATG9A could partially abolish miR-195-5p-mediated suppression of CC cells. Our findings improve our understanding of the mechanism of action of miR-195-5p in the malignant behaviour of CC. miR-195-5p is likely to be a promising cancer suppressor gene, which provides clinical evidence for targeted therapy of CC.

Colorectal cancer-secreted exosomal circ_001422 plays a role in regulating KDR expression and activating mTOR signaling in endothelial cells by targeting miR-195-5p.

BACKGROUND: As non-coding RNAs, exosomal circular RNAs (circRNAs) regulate colorectal cancer (CRC) progression, although the functional mechanisms by which such molecules affect the tumor microenvironment are still elusive. Herein, we aimed to explore the potential clinical significance of a signature of five serum-derived circRNAs in CRC and investigated the mechanisms underlying endothelial cell angiogenesis mediated by CRC-secreted exosomal circ_001422. METHODS: The expression of a signature of five serum-derived circRNAs (circ_0004771, circ_0101802, circ_0082333, circ_0072309, and circ_001422) were measured by RT-qPCR, and their associations with tumor staging and lymph node metastasis were further evaluated in CRC patients. In silico analysis was used to show the relationship between circ_001422, miR-195-5p, and KDR, validated by dual-luciferase reporter and Western blotting assays. CRC cell-derived exosomes were isolated and characterized by scanning electron microscopy and Western blotting. Endothelial cell uptake of PKH26-labeled exosomes was demonstrated using a spectral confocal microscope. In vitro genetic strategies were used to exogenously alter the expression level of circ_001422 and miR-195-5p expression. Cell proliferation assay, transwell migration assay, and capillary tube formation assay were conducted to explore the role of CRC-secreted exosomal circ_001422 in endothelial cell function in vitro. RESULTS: The expression levels of serum-derived circ_0004771, circ_0101802, circ_0082333, and circ_001422 were significantly higher in CRC and were positively correlated with the lymph node metastasis status. However, circ_0072309 showed a significant down-regulation in CRC than in healthy individuals. Furthermore, a higher expression level of circ_001422 in both cellular and exosomal fractions was found in HCT-116 CRC cells. We found that HCT-116 exosomes considerably enhanced proliferation and migration of endothelial cells through shuttling of circ_001422. We also observed that exosomes derived from HCT-116 cell, but not non-aggressive Caco-2 CRC cells, increased in vitro tubulogenesis of endothelial cells. Importantly, knockdown of circ_001422 impaired the capability of endothelial cells to form the capillary-like tube structures. CRC-secreted circ_001422 acted as an endogenous miR-195-5p sponge to inhibit miR-195-5p activity, which led to increased KDR expression and mTOR signaling activation in endothelial cells. Importantly, ectopic expression of miR-195-5p mimicked the effect of circ_001422 silencing on KDR/mTOR signaling in endothelial cells. CONCLUSION: This study attributed a biomarker role for circ_001422 in CRC diagnosis and proposed a novel mechanism whereby circ_001422 up-regulates KDR through sponging miR-195-5p. These interactions may give rise to the activation of mTOR signaling and may be a possible clarification for the pro-angiogenesis effects of CRC-secreted exosomal circ_001422 on endothelial cells.

Regulation of Schwann cell proliferation and migration via miR-195-5p-induced Crebl2 downregulation upon peripheral nerve damage.

Background: Schwann cells acquire a repair phenotype upon peripheral nerve injury (PNI), generating an optimal microenvironment that drives nerve repair. Multiple microRNAs (miRNAs) show differential expression in the damaged peripheral nerve, with critical regulatory functions in Schwann cell features. This study examined the time-dependent expression of miR-195-5p following PNI and demonstrated a marked dysregulation of miR-195-5p in the damaged sciatic nerve. Methods: CCK-8 and EdU assays were used to evaluate the effect of miR-195-5 on Schwann cell viability and proliferation. Schwann cell migration was tested using Transwell and wound healing assays. The miR-195-5p agomir injection experiment was used to evaluate the function of miR-195-5p in vivo. The potential regulators and effects of miR-195-5p were identified through bioinformatics evaluation. The relationship between miR-195-5p and its target was tested using double fluorescence reporter gene analysis. Results: In Schwann cells, high levels of miR-195-5p decreased viability and proliferation, while suppressed levels had the opposite effects. However, elevated miR-195-5p promoted Schwann cell migration determined by the Transwell and wound healing assays. In vivo injection of miR-195-5p agomir into rat sciatic nerves promote axon elongation after peripheral nerve injury by affecting Schwann cell distribution and myelin preservation. Bioinformatic assessment further revealed potential regulators and effectors for miR-195-5p, which were utilized to build a miR-195-5p-centered competing endogenous RNA network. Furthermore, miR-195-5p directly targeted cAMP response element binding protein-like 2 (Crebl2) mRNA via its 3'-untranslated region (3'-UTR) and downregulated Crebl2. Mechanistically, miR-195-5p modulated Schwann cell functions by repressing Crebl2. Conclusion: The above findings suggested a vital role for miR-195-5p/Crebl2 in the regulation of Schwann cell phenotype after sciatic nerve damage, which may contribute to peripheral nerve regeneration.

MiR-195-5p is involved in testicular ischemia/reperfusion injury by directly targeting PELP1 and regulating spermatogonia pyroptosis.

BACKGROUND AND OBJECTIVE: Ischemia/reperfusion injury (IRI), which is characterized by testicular torsion and causes permanent impairment of spermatogenic function, is linked with pyroptosis. Studies have implicated endogenous small non-coding RNAs in IRI development across various organs. In this study, we elucidated the mechanism underlying miR-195-5p's action in regulating pyroptosis in testicular IRI. METHODS: We established two models, namely a testicular torsion/ detorsion (T/D) mouse model and an oxygen-glucose deprivation/reperfusion (OGD/R)-treated germ cell model. Hematoxylin and eosin staining was performed to evaluate the testicular ischemic injury. The expression of pyroptosis-related proteins and reactive oxygen species production in testis tissues were detected using Western blotting, quantitative real-time PCR, malondialdehyde and superoxide dismutase assay kits and immunohistochemistry. Cell viability and cytotoxicity were evaluated using CCK-8 and LDH assays, whereas expression patterns of inflammatory proteins were measured using ELISA, immunofluorescence, and western blot assays. miR-195-5p interaction with PELP1 was validated by conducting the luciferase enzyme reporter test. RESULTS: Pyroptosis-related proteins NLRP3, GSDMD, IL-1ß, and IL-18 were significantly upregulated following testicular IRI. A similar pattern was observed in the OGD/R model. miR-195-5p was significantly downregulated in mouse IRI testis tissue and OGD/R-treated GC-1 cells. Notably, miR-195-5p downregulation promoted whereas its upregulation attenuated pyroptosis in OGD/R-treated GC-1 cells. Furthermore, we found that PELP1 is a miR-195-5p target. miR-195-5p attenuated pyroptosis in GC-1 cells by inhibiting PELP1 expression during OGD/R, and this protective effect was blocked upon miR-195-5p downregulation. Collectively, these results indicated that miR-195-5p inhibits testicular IRI-induced pyroptosis by targeting PELP1, suggesting that it has the potential to serve as a novel target for the future development of therapies for testicular torsion.

LncRNA FGD5-AS1 Promotes Abdominal Aortic Aneurysm Growth Through the Activation of MMP3 in Vascular Smooth Muscle Cells.

Long noncoding RNAs (lncRNAs) can serve as treatment targets for abdominal aortic aneurysms (AAAs). Nonetheless, the exact role of FGD5 antisense RNA 1 (FGD5-AS1) in AAAs is unclear. Therefore, this study investigated the contribution of FGD5-AS1 to AAA growth regulated by vascular smooth muscle cells (VSMCs) and its potential mechanisms. ApoE-/- mice were used to establish the angiotensin II (Ang II)-elicited AAA model. RNA pull-down assay and dual luciferase reporter assay (DLRA) in human VSMCs were used in examining the interactions between FGD5-AS1 and its downstream proteins or miRNA targets. FGD5-AS1 expression in the mouse Ang II perfusion group was dramatically increased relative to the PBS-infused group. In the mouse AAA model, FGD5-AS1 overexpression induced SMC apoptosis, thereby promoting AAA growth. miR-195-5p acts as a potential FGD5-AS1 downstream target, whereas FGD5-AS1 promotes MMP3 expression by inhibiting miR-195-5p expression, thereby inhibiting proliferation and promoting apoptosis of smooth muscle cells. LncRNA FGD5-AS1 is detrimental to the proliferation and survival of SMCs during AAA growth. Therefore, FGD5-AS1 could be a novel treatment target for AAA.

Extracellular Vesicle-Packaged miR-195-5p Sensitizes Melanoma to Targeted Therapy with Kinase Inhibitors.

Management of advanced melanoma remains challenging, with most BRAF (B-Raf proto-oncogene, serine/threonine kinase)-mutated metastatic patients relapsing within a few months upon MAPK inhibitors treatment. Modulation of tumor-derived extracellular vesicle (EVs) cargo with enrichment of antitumoral molecules is a promising strategy to impair tumor progression and increase treatment response. Herein, we report that restored expression of miR-195-5p, down-regulated in melanoma favoring drug resistance, increases the release of EVs enriched in the tumor suppressor miRNAs, miR-195-5p, miR-152-3p, and miR-202-3p. Incorporating these EVs by bystander tumor cells resulted in decreased proliferation and viability, accompanied by a reduction in CCND1 and YAP1 mRNA levels. Upon treatment with MAPK inhibitors, miR-195 EVs significantly decreased BCL2-L1 protein levels and increased cell death ratio and treatment efficacy. Additionally, EVs exogenously loaded with miR-195-5p by electroporation reduced tumor volume in vivo and impaired engraftment and growth of xenografts implanted with melanoma cells exposed to MAPK inhibitors. Our study shows that miR-195-5p antitumoral activity can be spread to bystander cells through EVs, improving melanoma response to targeted therapy and revealing a promising EV-based strategy to increase clinical response in patients harboring BRAF mutations.

Circular RNA circSEMA5A facilitates colorectal cancer development by regulating microRNA-195-5p to target CCNE1 axis.

Colorectal cancer (CRC) is one of the most prevalent malignancies with a high mortality rate worldwide. Circular RNAs (circRNAs) have lately emerged as key molecules involved in cancer development and metastasis. CircSEMA5 is reported to be oncogenic in some cancers, yet its role in the pathogenesis of CRC remains unknown. Herein, we attempted to investigate the functional role and molecular mechanism of circSEMA5A underlying CRC progression. RT-qPCR and RNase R digestion assays were used to evaluate circSEMA5A expression characteristics in CRC cells. Loss-of-function assays were performed to clarify circSEMA5A role in CRC biological processes. Bioinformatics and mechanism experiments were conducted to assess the association of circSEMA5A or CCNE1 with miR-195-5p in CRC cells. Rescue assays were conducted to explore the regulatory function of circSEMA5A-miR-195-5p-CCNE1 in CRC cellular processes. Through bioinformatics and functional screening, we found that circSEMA5A was highly expressed in CRC cells and was mainly localized in the nucleus. CircSEMA5A promoted CRC proliferative, migratory, and invasive capabilities in cultured cells and facilitated the tumorigenic process in xenografts; however, circSEMA5A silencing repressed tumor metastasis in CRC cells. Mechanistically, circSEMA5A was competitively bound with miR-195-5p to upregulate CCNE1 expression. Moreover, the impact of circSEMA5A knockdown on CRC cell proliferative, migratory, and invasive capabilities was countervailed by miR-195-5p inhibitor or CCNE1 overexpression. To summarize, circSEMA5A is a novel circRNA that serves as an oncogene in CRC progression. CircSEMA5A facilitates CRC cell malignancy and tumor growth through sponging miR-195-5p to upregulate CCNE1, thus providing a new direction for CRC diagnosis and targeted therapy.

LncRNA CERS6-AS1 Is a Tumor Promoter in Cervical Cancer by Sponging miR-195-5p.

OBJECTIVE: CERS6 antisense RNA 1 (CERS6-AS1), a long non-coding RNA (lncRNA), plays a role in the malignant progression of a variety of cancers. However, it is unclear whether it affects the malignant behavior of cervical cancer (CC) cells. METHODS: CERS6-AS1 and miR-195-5p expression was estimated in CC via qRT-PCR. CCK-8, caspase-3 activity, scratch, and Transwell assays were performed to detect CC cell viability, caspase-3 activity, migration, and invasion in vitro. A tumor xenograft experiment was designed to study the growth of CC tumors in vivo. RIP and luciferase reporter experiments verified the relationship between CERS6-AS1 and miR-195-5p. RESULTS: CERS6-AS1 overexpression and poor miR-195-5p levels were observed in CC. Inhibition of CERS6-AS1 impaired the viability, invasion, and migration of CC cells, promoted apoptosis, and suppressed tumor growth. In terms of the underlying mechanism, CERS6-AS1, as a competitive endogenous RNA (ceRNA), participated in the regulation of miR-195-5p levels in CC cells. Functionally, miR-195-5p interference attenuated the inhibitory effect of CERS6-AS1 on the malignant behaviors of CC cells. CONCLUSION: CERS6-AS1 acts as an oncogene in CC, in vivo and in vitro, by negatively regulating miR-195-5p.

Exosomal transfer of miR-195-5p restrains lung adenocarcinoma progression.

Exosome is an important way for tumor cells to communicate with other cells and plays an important role in tumor progression. Previous studies revealed that miR-195-5p acts as a tumor suppressor in lung cancer. However, the role and molecular mechanism of exosomal transferred miR-195-5p in lung adenocarcinoma (LAC) remains unknown. Here, we found that miR-195-5p expression in circulating exosomes of LAC patients was lower than that of healthy controls. Meanwhile, the expression of exosomal miR-195-5p from normal bronchial epithelial cell line BEAS-2B cells was significantly higher than that of lung cancer cell lines. The exosome labeling assay confirmed that BEAS-2B cells-derived exosomes could be captured by lung cancer cells. Furthermore, exosomal miR-195-5p derived from BEAS-2B cells remarkably inhibited the proliferation, migration, invasion of lung cancer cells, and tumor growth in vivo. In addition, exosomal miR-195-5p from BEAS-2B cells also suppressed the tube-forming ability of vascular endothelial cells. Moreover, we verified that miR-195-5p decreased apelin (APLN) expression to inactivate the Wnt signaling pathway, thereby inhibiting tumor invasiveness and angiogenesis. In conclusion, our research shows that exosomal miR-195-5p from normal bronchial epithelial cells hinders the progression of LAC, suggesting that regulation of exosomal miR-195-5p provides a novel strategy for LAC treatment.

Hsa-miR-195-5p Inhibits Autophagy and Gemcitabine Resistance of Lung Adenocarcinoma Cells via E2F7/CEP55.

Lung adenocarcinoma (LUAD) is a common malignancy. Many studies have shown that LUAD is resistant to gemcitabine chemotherapy, resulting in poor treatment outcomes in patients. We designed this study to reveal influences of hsa-miR-195-5p/E2F7/CEP55 axis on gemcitabine resistance and autophagy of LUAD cells. The expression data of LUAD-related mRNAs were downloaded from TCGA-LUAD database for differential expression analysis. The bioinformatics databases (hTFtarget, starBase and TargetScan) were used to predict the upstream and downstream regulatory molecules of E2F7. Then the binding relationships between E2F7 and regulatory molecules were verified by ChIP and dual-luciferase reporter assay. qRT-PCR and western blot were used to detect the mRNA and protein levels of has-miR-195-5p, E2F7, and CEP55. CCK-8 assay was used to analyze the half-maximal inhibitory concentration (IC50) and cell proliferation ability of LUAD cells after gemcitabine treatment. Apoptosis was detected by flow cytometry. Apoptosis/autophagy markers and LC3 aggregation were detected by western blot and immunofluorescence, respectively. Finally, the mouse transplantation model was constructed to verify the regulation mechanism in vivo. In LUAD cells and tissues, E2F7 and CEP55 were highly expressed, while has-miR-195-5p was relatively less expressed. The ChIP or dual-luciferase assays demonstrated the binding relationships of E2F7 to the CEP55 promoter region and has-miR-195-5p to the 3'-UTR of E2F7. Cell experiments demonstrated that overexpression of hsa-miR-195-5p stimulated LUAD cell apoptosis and inhibited autophagy and gemcitabine resistance, while further overexpression E2F7/CEP55 could reverse the impact by hsa-miR-195-5p overexpression. In vivo experiments identified that hsa-miR-195-5p/E2F7/CEP55 axis constrained the growth of LUAD tumor. Hsa-miR-195-5p promoted apoptosis, repressed proliferation, and autophagy via E2F7/CEP55 and reduced gemcitabine resistance in LUAD, indicating that hsa-miR-195-5p/E2F7/CEP55 may be a novel target for LUAD.