Indocyanine green based photodynamic therapy for keloids: Fundamental investigation and clinical improvement.

BACKGROUND: Keloid, a prevalent pathological skin lesion, presents significant challenges in terms of treatment efficacy. Photodynamic therapy (PDT), an increasingly popular adjuvant treatment, has shown significant potential in the management of various disorders, including cancer. However, the therapeutic potential of indocyanine green-mediated photodynamic therapy (ICG-PDT) for keloids has not yet been demonstrated. METHODS: In this study, we divided the experimental groups into control group, Photothermal Therapy group, Photodynamic Therapy group, and Combined Therapy group. The in vitro investigation aimed to optimize the clinical application of PDT for keloid treatment by elucidating its underlying mechanism. Subsequently, on this basis, we endeavored to manage a clinical case of keloid by employing surgical intervention in conjunction with modified ICG-PDT. RESULTS: Our investigation revealed an unexpected outcome that ICG-PDT maximally inhibited the cellular activity and migration of keloid fibroblasts only when photodynamic mechanism took effect. Additionally, the induction of autophagy and apoptosis, as well as the inhibition of collagen synthesis, were particularly evident in this experimental group. Furthermore, the above therapeutic effect could be achieved at remarkably low drug concentrations. Building upon the aforementioned experimental findings, we successfully optimized the treatment modality for the latest case and obtained a more favorable treatment outcome. CONCLUSIONS: This study investigated the mechanism of ICG-PDT treatment and optimized the in vivo treatment regimen, demonstrating the significant therapeutic potential of ICG-PDT treatment in clinical keloid treatment.

HPV E6 promotes cell proliferation of cervical cancer cell by accelerating accumulation of RBM15 dependently of autophagy inhibition.

The mechanism of m6A modification in HPV-related cervical cancer remains unclear. This study explored the role of methyltransferase components in HPV-related cervical cancer and the mechanism. The levels of methyltransferase components and autophagy, ubiquitylation of RBM15 protein and the co-localization of lysosomal markers LAMP2A and RBM15 were measured. CCK-8 assay, flow cytometry, clone formation experiment and immunofluorescence assay were conducted to measure cell proliferation. The mouse tumor model was developed to study the cell growth in vivo. The binding of RBM15 to c-myc mRNA and m6A modifcation of c-myc mRNA were analyzed. The expressions of METTL3, RBM15 and WTAP were higher in HPV-positive cervical cancer cell lines than those in HPV-negative cells, especially RBM15. HPV-E6 knock-down inhibited the expression of RBM15 protein and promoted its degradation, but couldn't change its mRNA level. Autophagy inhibitor and proteasome inhibitor could reverse those effects. HPV-E6 siRNA could not enhance ubiquitylation modification of RBM15, but could enhance autophagy and the co-localization of RBM15 and LAMP2A. RBM15 overexpression could enhance cell proliferation, block the inhibitory effects of HPV-E6 siRNA on cell growth, and these effects could be reserved by cycloeucine. RBM15 could bind to c-myc mRNA, resulting in an increase to m6A level and protein expression of c-myc, which could be blocked by cycloeucine. HPV-E6 can downregulate autophagy, inhibit the degradation of RBM15 protein, induce the accumulation of intracellular RBM15, and increase the m6A modification on c-myc mRNA, resulting in an increase of c-myc protein and a growth promotion for cervical cancer cells.

LINC01063 functions as an oncogene in melanoma through regulation of miR-5194-mediated SOX12 expression.

Melanoma is one of the most aggressive skin cancers and a major cause of cancer-linked deaths worldwide. As the morbidity and mortality of melanoma are increasing, it is necessary to elucidate the potential mechanism influencing melanoma progression. Tumor tissues and adjacent normal tissues (5 cm away from tumors) from 22 melanoma patients at the I-II stage and 39 patients at the III-VI stage were acquired. The expression of LINC01063 in melanoma was estimated by quantitative PCR. Functional assays were employed to investigate the function of LINC01063 in melanoma. Mechanism assays were adopted to explore the mechanism of LINC01063. LINC01063 knockdown impeded melanoma cell proliferation, migration, invasion, and epithelial-mesenchymal transition as well as melanoma tumor growth. Mechanistically, LINC01063 acted as an miR-5194 sponge to upregulate SOX12 expression. Finally, LINC01063 was tested to facilitate the malignant behaviors of melanoma cells via targeting miR-5194/SOX12. LINC01063 was significantly upregulated in melanoma. Specifically, LINC01063 displayed a higher level in patients at an advanced stage or with metastasis than those at an early stage or without metastasis. Our study revealed the oncogenic effects of LINC01063 on melanoma cell/tumor growth and its molecular mechanism involving miR-5194/SOX12, which might support LINC01063 to be the potential prognostic or therapeutic biomarker against melanoma.

Circular RNA circLMO1 Suppresses Cervical Cancer Growth and Metastasis by Triggering miR-4291/ACSL4-Mediated Ferroptosis.

Background: A number of studies have demonstrated that circular RNA (circRNA) plays a critical role in tumorigenesis and tumor progression. However, the biological effects of most circRNAs on cervical cancer remain unclear. Hsa_circ_0021087 (thereafter named circLMO1) is a circRNA generated from the circularization of exon 2 and exon 3 of LIM Domain Only 1 (LMO1) and first identified as a tumor suppressor in gastric cancer. We aimed to identify the role of circLMO1 in cervical cancer progression. Methods: CircLMO1 was verified through qPCR and Sanger sequencing. The biological role of circLMO1 in regulating cervical cancer growth and metastasis was investigated both in vitro and in the nude mouse xenograft tumor model. The dual luciferase reporter assay and rescue experiment were conducted to evaluate the interactions among circLMO1, microRNA (miR)-4291, and acyl-CoA synthetase long chain family member 4 (ACSL4). The role of circLMO1 in regulating ferroptosis was assessed by analyzing lipid reactive oxygen species (ROS), and malonyl dialdehyde (MDA), and glutathione (GSH) content. Results: The level of circLMO1 was down-regulated in cervical cancer tissues and was associated with the International Federation of Gynecology and Obstetrics (FIGO) staging. Functionally, circLMO1 overexpression inhibited cervical cancer growth and metastasis both in vitro and in vivo, whereas circLMO1 depletion promoted cervical cancer cell proliferation and invasion. Mechanistically, circLMO1 acted as a competing endogenous RNA (ceRNA) by sponging miR-4192 to repress target gene ACSL4. CircLMO1 promoted cervical cancer cell ferroptosis through up-regulating ACSL4 expression. Overexpression of miR-4291 or knockdown of ACSL4 reversed the effect of circLMO1 on facilitating ferroptosis and repressing cervical cancer cell proliferation and invasion. Conclusion: CircLMO1 acted as a tumor suppressor of cervical cancer by regulating miR-4291/ACSL4-mediated ferroptosis, and could be a promising biomarker for the clinical management of cervical cancer.

Cinnamaldehyde inhibits psoriasis­like inflammation by suppressing proliferation and inflammatory response of keratinocytes via inhibition of NF­κB and JNK signaling pathways.

Psoriasis is a systemic immune­mediated inflammatory disease characterized by uncontrolled keratinocyte proliferation and poor differentiation. Cinnamaldehyde (CIN) has been shown to inhibit the proliferation and inflammatory response of primary and immortalized immune cells. However, to the best of our knowledge, the role of CIN in the progression of psoriasis remains unclear. Therefore, the present study aimed to investigate the biological role of CIN in psoriasis. To mimic abnormal proliferation and differentiation in keratinocytes in vitro, normal human epidermal keratinocytes (NHEKs) were stimulated with M5 (IL­1α, IL­17A, IL­22, oncostatin M and TNF­α). The viability and proliferation of NHEKs were analyzed using Cell Counting Kit­8 and 5­Ethynyl­2'­deoxyuridine assays, respectively. Western blotting was used to analyze the expression levels of keratin 1, filaggrin and loricrin in NHEKs. The results of the present study revealed that CIN significantly inhibited the proliferation and cell cycle progression, and promoted the differentiation of M5­stimulated NHEKs. CIN also markedly attenuated the extent of oxidative stress­induced damage in M5­stimulated NHEKs. Moreover, CIN ameliorated M5­induced inflammatory injury in NHEKs, as evidenced by the decreased levels of multiple inflammatory factors. Furthermore, CIN notably downregulated the expression levels of phosphorylated (p)­inhibitor of NF­κB, p­p65 and p­JNK in M5­stimulated NHEKs. In conclusion, the present data suggested that CIN may protect NHEKs against M5­induced hyperproliferation and inflammatory injury via inhibition of NF­κB and JNK signaling pathways. These results provide a novel insight on the role of CIN in psoriasis.

Curcumol may alleviate psoriasis-like inflammation by inhibiting keratinocyte proliferation and inflammatory gene expression via JAK1/STAT3 signaling.

Psoriasis is a chronic inflammatory skin disease characterized by abnormal proliferation and differentiation of keratinocytes. Since curcumol exhibits anti-inflammatory properties in various diseases, we investigated its anti-inflammatory potential in stimulated human keratinocytes. Our data show that curcumol significantly inhibits proliferation and induces cell cycle arrest in NHEK cells stimulated with proinflammatory cytokines (IL-1α, IL-17A, IL-22, oncostatin M, and TNF-α; mix M5). In addition, curcumol markedly ameliorates inflammatory response and promotes differentiation of M5-stimulated NHEK cells. Curcumol inhibits activity of JAK1, resulting in the inhibition of STAT3, downregulation of cyclin D2, and cell cycle arrest in stimulated NHEK cells. Together, our data show that curcumol reduces proliferation and inflammatory gene expression in stimulated keratinocytes by inhibiting the JAK1/STAT3 signaling, suggesting that it might serve as a potential therapeutic option for the treatment of psoriasis.

FNDC3B and BPGM Are Involved in Human Papillomavirus-Mediated Carcinogenesis of Cervical Cancer.

Human papillomavirus (HPV)-mediated cervical carcinogenesis is a multistep progressing from persistent infection, precancerous lesion to cervical cancer (CCa). Although molecular alterations driven by viral oncoproteins are necessary in cervical carcinogenesis, the key regulators behind the multistep process remain not well understood. It is pivotal to identify the key genes involved in the process for early diagnosis and treatment of this disease. Here we analyzed the mRNA expression profiles in cervical samples including normal, cervical intraepithelial neoplasia (CIN), and CCa. A co-expression network was constructed using weighted gene co-expression network analysis (WGCNA) to reveal the crucial modules in the dynamic process from HPV infection to CCa development. Furthermore, the differentially expressed genes (DEGs) that could distinguish all stages of progression of CCa were screened. The key genes involved in HPV-CCa were identified. It was found that the genes involved in DNA replication/repair and cell cycle were upregulated in CIN compared with normal control, and sustained in CCa, accompanied by substantial metabolic shifts. We found that upregulated fibronectin type III domain-containing 3B (FNDC3B) and downregulated bisphosphoglycerate mutase (BPGM) could differentiate all stages of CCa progression. In patients with CCa, a higher expression of FNDC3B or lower expression of BPGM was closely correlated with a shorter overall survival (OS) and disease-free survival (DFS). A receiver operating characteristic (ROC) analysis of CIN and CCa showed that FNDC3B had the highest sensitivity and specificity for predicting CCa development. Taken together, the current data showed that FNDC3B and BPGM were key genes involved in HPV-mediated transformation from normal epithelium to precancerous lesions and CCa.

HPV16 E6 oncoprotein-induced upregulation of lncRNA GABPB1-AS1 facilitates cervical cancer progression by regulating miR-519e-5p/Notch2 axis.

Human papillomaviruses 16 (HPV16) is the primary causative agent of cervical cancer (CC). E6 oncoprotein plays a crucial role in cervical carcinogenesis and commonly cause the dysregulation of the long noncoding RNAs (lncRNAs) expression. However, the biological function of lncRNAs in HPV16-related CC remains largely unexplored. In the present study HPV16 E6-induced differential expression of lncRNAs, miRNA, and mRNA were identified using microarray-based analysis and verified in tumor r cell lines and tumor tissues, and the function of lncRNA in CC was investigated in vitro and in vivo. We found that an lncRNA, named GABPB1-AS1, was significantly upregulated in HPV16-positive CC tissues and cell lines. GABPB1-AS1 expression in HPV16-positive CC tissues was positively associated with tumor size, lymph node metastasis, and FIGO stage. High expression of GABPB1-AS1 was correlated with a poor prognosis for HPV16-positive CC patients. Functionally, E6-induced GABPB1-AS1 overexpression facilitated CC cells proliferation and invasion in vitro and in vivo. Mechanistically, GABPB1-AS1 acted as a competing endogenous RNA (ceRNA) by sponging miR-519e-5p, resulting in the de-repression of its target gene Notch2 which is well known as an oncogene. Therefore, GABPB1-AS1 functioned as a tumor activator in CC pathogenesis by binding to miR-519e-5p and destroying its tumor suppressive function. Collectively, current results demonstrate that GABPB1-AS1 is associated with CC progression, and may be a promising biomarker or target for the clinical management of CC.

Long Noncoding RNA CAR10 Contributes to Melanoma Progression By Suppressing miR-125b-5p to Induce RAB3D Expression.

BACKGROUND: Melanoma is a very malignant skin cancer with high mortality and unsatisfactory prognosis. Many long noncoding RNAs (lncRNAs) have been reported to be aberrantly expressed in melanoma. How lncRNA regulates melanoma progression is poorly defined. LncRNA CAR10 has been shown to regulate the progression of several cancers and its role in melanoma remains unclear. This study aims to determine the role and mechanism of lncRNA CAR10 in the regulation of melanoma progression. METHODS: qRT-PCR was utilized to analyze CAR10 in melanoma human tissues and cell lines while Kaplan-Meier curve was used to examine the survival rate. CCK8 assay and EdU assay were used to assess cell proliferation when Transwell assay was conducted to determine migration and invasion. And tumor xenograft assay was performed to evaluate tumor growth in vivo. Additionally, luciferase assay and RNA pulldown assay were performed to analyze the interactions among CAR10, miR-125b-5p and RAB3D. RESULTS: LncRNA CAR10 was upregulated in melanoma tissues and cell lines. Upregulation of CAR10 predicted a poor prognosis in patients with melanoma. CAR10 knockdown suppressed proliferation, migration and invasion of melanoma cells in vitro. CAR10 silencing attenuated tumor growth in vivo. CAR10 inhibited miR-125b-5p activity to upregulate RAB3D expression. And miR-125b-5p/RAB3D signaling is crucial for CAR10-dependent melanoma progression. CONCLUSION: Our work suggests that lncRNA CAR10 promotes melanoma growth and metastasis through modulating miR-125b-5p/RAB3D axis.

MicroRNA­155 inhibits the proliferation of CD8+ T cells via upregulating regulatory T cells in vitiligo.

It has been reported that loss and degradation of epidermal melanocytes is closely associated with the pathogenesis of vitiligo. In addition, CD8+ T and regulatory T (Treg) cells serve an important role during these two processes. MicroRNA­155 (miR­155) is known to contribute to the pathogenesis of vitiligo; however, the mechanism by which miR­155 regulates the development of vitiligo remains unclear. In the present study, naïve T and CD8+ T cells were isolated from a patient with non­segmental vitiligo by flow cytometry. The cells were differentiated into Treg cells by treatment with interleukin­2, transforming growth factor­ß and retinoic acid. In addition, miR­155 agonists and antagonists were used to investigate the effect of miR­155 on the proliferation of CD8+ T cells, Treg cells and melanocytes. The results demonstrated that the miR­155 agonist significantly decreased the rate of CD8+ T cell growth, as well as promoted the proliferation of melanocytes by inducing an increase in the percentage of Treg cells. By contrast, the miR­155 antagonist inhibited the proliferation of melanocytes by decreasing the percentage of Treg cells. miR­155 protected melanocyte survival by increasing the number of Treg cells and by decreasing the number of CD8+ T cells. Therefore, these data may provide a new prospect for the treatment of vitiligo.

MicroRNA-664 suppresses the growth of cervical cancer cells via targeting c-Kit.

BACKGROUND: Cervical cancer is the second most common malignant cancer in women worldwide. Evidence indicated that miR-664 was significantly downregulated in cervical cancer. However, the mechanisms by which miR-664 regulates the tumorigenesis of cervical cancer remain unclear. Thus, this study aimed to investigate the role of miR-664 in cervical cancer. METHODS: Quantitative reverse transcription polymerase chain reaction was used to detect the level of miR-664 in tumor tissues and cell line. The dual luciferase reporter system assay and Western blotting were used to explore the interaction of miR-664 and c-Kit in cervical cancer. RESULTS: The expression of miR-664 in patients with cervical cancer was dramatically decreased compared with that in adjacent tissues. MiR-664 mimics significantly inhibited proliferation in SiHa cells via inducing apoptosis. In addition, miR-664 mimics induced apoptosis in SiHa cells via increasing the expressions of Bax and active caspase 3 and decreasing the level of Bcl-2. Moreover, dual-luciferase assay showed that c-Kit was the directly binding target of miR-664 in SiHa cells; overexpression of miR-664 downregulated the expression of c-Kit. Meanwhile, upregulation of miR-664 significantly decreased the levels of c-Myc and Cyclin D in cells. Furthermore, miR-664 markedly inhibited tumor growth of cervical cancer in xenograft. CONCLUSION: Our data indicated that miR-664 exerted antitumor effects on SiHa cells by directly targeting c-Kit in vitro and in vivo. Therefore, miR-664 might be a potential therapeutic target for the treatment of patients with cervical cancer.

Poly(I:C) enhanced anti-cervical cancer immunities induced by dendritic cells-derived exosomes.

Dendritic cell (DC)-derived exosomes (Dexo) has been confirmed to be able to induce the specific anti-tumor immune response ex vivo and in vivo. Here, the aim of this study was to evaluate the application of the antigen-pulsed Dexo as a new vaccination platform in immunotherapy for cervical cancer. The immunogenic profile of the different Dexo was assessed by the cell proliferation, cytokines secretion and effector functions of CD8+ T cells and the splenocytes from Dexo-vaccinated mice. Furthermore, the anti-tumor immunity elicited by Dexo was further compared in cervical cancer-bearing mice. Dexo from DCs loaded with E749-57 peptide could efficiently induce the cytotoxic activity of CD8+ T cells on TC-1 tumor cells ex vivo, the proliferation and IFN-γ excretion of CD8+ T cells. Moreover, Dexo vaccine promoted the immune responses of vaccinated mice splenocytes induced by antigen E7 in vitro restimulation. Of note, poly(I:C) was significantly more potent inducer of the antigen-loaded Dexo mediated protective immunity responses for cervical cancer and further evidenced by that Dexo(E7+pIC) markedly inhibited the tumor growth and improved the survival rate of the tumor-bearing mice. We provided evidence that poly(I:C) dramatically increased the potent antitumoral immunity induced by antigen-pulsed Dexo for ameliorating cervical cancer.

Neutrophil depletion improves diet-induced non-alcoholic fatty liver disease in mice.

PURPOSE: Non-alcoholic fatty liver disease is highly associated with morbidity and mortality in population. Although studies have already demonstrated that the immune response plays a pivotal role in the development of non-alcoholic fatty liver disease, the comprehensive regulation is unclear. Therefore, present study was carried out to investigate the non-alcoholic fatty liver disease development under neutrophil depletion. METHODS: To achieve the aim of the study, C57BL/6 J mice were fed with high fat diet for 6 weeks before treated with neutrophil deplete antibody 1A8 or isotype control (200 µg/ mouse every week) for another 4 weeks. RESULTS: Treated with 1A8 antibody, obese mice exhibited better whole body metabolic parameters, including reduction of body weight gain and fasting blood glucose levels. Neutrophil depletion also effectively reduced hepatic structural disorders, dysfunction and lipid accumulation. Lipid ß-oxidative markers, phosphorylated-AMP-activated protein kinase α and phosphorylated-acetyl-CoA carboxylase levels were increased in 1A8 antibody-treated obese mouse group. The mitochondrial number and function were also reversed after 1A8 antibody treatment, including increased mitochondrial number, reduced lipid oxidative damage and enhanced mitochondrial activity. Furthermore, the expression of inflammatory cytokines, tumor necrosis factor-α, interleukin-6, and monocyte chemoattractant protein-1 were obviously reduced after neutrophil depletion, accompanied with decreased F4/80 mRNA level and macrophage percentage in liver. The decreased NF-κB signaling activity was also involved in the beneficial effect of neutrophil depletion. CONCLUSION: Taken together, neutrophil depletion could attenuate metabolic syndromes and hepatic dysfunction.

Downregulation of pyrroline-5-carboxylate reductase-2 induces the autophagy of melanoma cells via AMPK/mTOR pathway.

Melanoma is the most aggressive form of skin cancer and causes 50,000 deaths annually worldwide. The roles of proline-dependent process and autophagy have both been reported in studies on melanoma. In the present study, we focused on the effect of pyrroline-5-carboxylate reductase-2 (PYCR2) on inducing autophagy process in melanoma. The expression of PYCR2 was regulated by an RNAi technique, and the cell proliferation of A375 cell line was determined by methyl thiazolyl tetrazolium test; the effect of PYCR2 on the apoptosis process and AMPK/mTOR pathway was evaluated by flow cytometry assay and Western blot. It was found that silence of PYCR2 resulted in the decrease of proliferative ability and activation of AMPK/mTOR-induced autophagy of A375 cells. PYCR2 silencing also activated AMPK/mTOR pathway in another melanoma cell line, CHL-1. However, the overexpression of PYCR2 seemed to make no difference to the cell viability and targeted pathway. Our results offered a preliminary illustration on the mechanism of the PYCR2-dependent autophagy and showed that PYCR2 was a potential therapeutic target of melanoma.