Histology and histopathology Vol.39, nº2 (2024)
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- PublicationOpen AccessTRIM3 inhibits colorectal cancer cell migration and lipid droplet formation by promoting FABP4 degradation(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Zuo, Qi; Xu, Qimei; Li, Zhen; Luo, Dixian; Peng, Hanwu; Duan, ZhiThis study is to investigate the regulation of TRIM3/FABP4 on colorectal cancer (CRC) cell migration and lipid metabolism. After transfection of HCT116, LoVo, or SW480 cells, the expression of FABP4, TRIM3, N-cadherin, Vimentin, E-cadherin, and lipid droplet (LD) formation-related genes was measured by qRT-PCR or western blot assays. Wound healing and Transwell assays were applied to detect CRC cell migration and invasion abilities. The levels of triglyceride (TG) and total cholesterol (TC) were measured and the formation of LDs was observed. Additionally, the relationship between FABP4 and TRIM3 was confirmed by Co-IP and ubiquitination assays. Furthermore, a liver metastasis model of CRC was established to explore the effect of FABP4 on CRC tumor metastasis in vivo. FABP4 was upregulated in CRC cells. Downregulation of FABP4 or upregulation of TRIM3 resulted in repressed cell migration and invasion, decreased TG and TC levels, and reduced numbers of LDs. In nude mice, knockdown of FABP4 reduced metastatic nodules in the liver. Mechanistically, TRIM3 combined FABP4 and decreased its protein expression by ubiquitination. Overexpressed FABP4 reversed the influence of TRIM3 upregulation on CRC cell migration and LD formation. In conclusion, underexpressed TRIM3 suppressed FABP4 ubiquitination and accelerated CRC cell migration and LD formation
- PublicationOpen AccessInhibition of ubiquitin specific peptidase 8 is effective against 5-fluorouracil resistance in colon cancer via suppressing EGFR and EGFR-mediated signaling pathways(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Xi, Changlei; Gong, Zhilin; Ye, Hui; Cao, Longlei; Yu, JieBackground. The identification of a sensitizing strategy to overcome 5-fluorouracil (5-FU) therapeutic resistance is needed in colon cancer. Recent studies highlight the oncogenic role of ubiquitin specific peptidase 8 (USP8) in many cancers. In line with these efforts, this work investigated the therapeutic potential of targeting USP8 in colon cancer. Methods. Immunohistochemistry was performed to determine USP8 expression level in colon cancer tissues and their adjacent normal tissues. Gain-of-function analysis via plasmid overexpression and loss-of-function analysis via siRNA knockdown were applied on cellular assays. The combinatory effects of USP8 inhibitor and cisplatin were determined using a colon xenograft mouse model. Immunoblotting was performed to investigate the molecular mechanism of USP8 inhibition in colon cancer cells. Results. Compared to normal counterparts, we showed that USP8 protein level was significantly higher in colon cancer tissues and cells. In addition, USP8 expression was not affected by prolonged exposure of colon cancer cells to 5-FU. USP8 was important for colon cancer cell growth and survival but not migration as assessed by loss-of-function and gain-of-function approaches. Pharmacological inhibition of USP8 using USP8 inhibitor is active against both sensitive and 5-FUresistant colon cancer cells. Of note, USP8 inhibitor significantly inhibited colon cancer formation and growth, and augmented in vivo efficacy of 5-FU without causing toxicity in mice. Mechanistic studies showed that USP8 inhibitor acted on colon cancer cells through suppressing EGFR and EGFR-mediated signalling pathways. Conclusions. Our work is the first to reveal the essential role of USP8 in colon cancer via EGFR oncogenic signalling pathways. Our findings provide a proof-of-concept that USP8 inhibitors are promising candidates to overcome 5-FU resistance in colon cancer
- PublicationOpen AccessCircRTTN upregulates EPHA2 to aggravate the malignant process of melanoma via sponging miR-890(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Wang, Yaqin; Gong, Junzuo; Ding, Xiaojie; Luo, ShuBackground. Malignant melanoma is a kind of tumor derived from melanocytes, which has the characteristics of drug resistance and distant metastasis. Accumulating evidence has demonstrated that circular RNAs (circRNAs) are involved in the pathogenesis of melanoma. Our current study aimed to investigate the role and mechanism of circRTTN in melanoma progression. Methods. The levels of circRTTN, microRNA-890 (miR-890) and EPH receptor A2 (EPHA2) were examined via quantitative real-time PCR (qRT-PCR) and Western blot. Cell Counting Kit-8 (CCK-8), colony formation, 5-Ethynyl-2’-deoxyuridine (EdU) staining, flow cytometry, transwell and tube formation assays were conducted to estimate the effects of circRTTN on growth, apoptosis, migration, invasion and angiogenesis of melanoma cells. Western blot was used to measure related marker protein levels. The interaction between miR-890 and circRTTN or EPHA2 was predicted by bioinformatics analysis and verified by dual-luciferase reporter and RNA Immunoprecipitation (RIP) assays. Xenograft assay was used to assess the effect of circRTTN in vivo. Results. CircRTTN and EPHA2 levels were upregulated, while miR-890 was down-regulated in melanoma tissues and cells. CircRTTN knockdown restrained cell proliferation, migration, invasion and angiogenesis, but promoted cell apoptosis in vitro. CircRTTN was an effective molecular sponge for miR890, and negatively regulated miR-890 expression. The suppressive role of circRTTN knockdown on cell growth, metastasis and angiogenesis in vitro was abated by blocking miR-890. MiR-890 directly targeted EPHA2. MiR-890 overexpression elicited a similar antitumor role in melanoma cells, which was abrogated by overexpression of EPHA2. In addition circRTTN knowdown markedly attenuated xenograft tumor growth in vivo. Conclusion. Our findings demonstrated that circRTTN mediated melanoma progression via regulating the miR-890/ EPHA2 axis.
- PublicationOpen AccessCalpain-specific breakdown fragment in human drusen(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Orihara, Kana; Kobayashi Otsugu, Momoko; Nakajima, Emi; Walkup, Ryan D.; Wilson, David J.; Shearer, Thomas R.; Azuma, MitsuyoshiPurpose. With aging and age-related macular dystrophy (AMD), proteolytic fragments are deposited in extracellular drusen located between the RPE and Bruch’s membrane. Localized hypoxia may be a risk factor for AMD. Our hypothesis is that following hypoxia, activation of proteolytic enzymes called calpains may cause proteolysis/degeneration of retinal cells and RPE. No direct evidence has yet demonstrated activation of calpains in AMD. The purpose of the present study was to identify calpain-cleaved proteins in drusen. Methods. Seventy-six (76) drusen were analyzed in human eye sections from six normal and twelve AMD human donor eyes. The sections were subjected to immunofluorescence for the calpain-specific 150 kDa breakdown product from α-spectrin, SBDP150-a marker for calpain activation, and for recoverin-a marker for photoreceptor cells. Results. Among 29 nodular drusen, 80% from normal eyes and 90% from AMD eyes stained positive for SBDP150. Among 47 soft drusen, mostly from AMD eyes, 72% stained positive for SBDP150. Thus, the majority of both soft and nodular drusen from AMD donors contained SBDP150. Conclusions. SBDP150 was detected for the first time in soft and nodular drusen from human donors. Our results suggest that calpain-induced proteolysis participates in the degeneration of photoreceptors and/or RPE cells during aging and AMD. Calpain inhibitors may ameliorate AMD progression
- PublicationOpen AccessEpigenetic upregulation of MNAT1 by SMYD2 is linked to PI3K/AKT activation and tumorigenesis of pancreatic adenocarcinoma(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Xu, Zhen; Liu, Yan; Pan, Zhi; Qin, LeiDysregulation of histone methyltransferase SET and MYND domain-containing protein 2 (SMYD2) has been correlated with human developmental disorders and cancers. This research aims to investigate the roles of SMYD2 and its interacted molecules in pancreatic adenocarcinoma (PAAD). Two PAAD-related gene expression datasets were downloaded to screen key molecules involved in tumor progression. SMYD2 was expressed at high levels in PAAD tissues and cells. SMYD2 silencing suppressed while its overexpression promoted proliferation, invasiveness, migration, apoptosis resistance, and cell cycle progression of PAAD cells. Target molecules of SMYD2 were predicted by online tools and validated by chromatin immunoprecipitation and luciferase assays. SMYD2 catalyzed H3K36me2 modification at the promoter region of MNAT1 component of CDK activating kinase (MNAT1) to promote its transcription. MNAT1 was correlated with an unfavorable clinical outcome of PAAD patients. Alteration of MNAT1 alone also affected the malignant behavior of PAAD cells. Moreover, MNAT1 overexpression in cells rescued the malignant phenotype of cells suppressed by SMYD2 silencing. MNAT1 activated the phosphatidyl inositol 3-kinase/protein kinase B (PI3K/AKT) signaling. In vivo, SMYD2 silencing decreased the growth rate and weight of xenograft tumors in nude mice. Overall, this paper demonstrates that SMYD2-mediated MNAT1 upregulation is linked to PAAD tumorigenesis via PI3K/ AKT pathway activation.
- PublicationOpen AccessUpregulated miR-125b mitigates inflammation, astrocyte activation, and dysfunction of spinal cord injury by inactivating the MAPK pathway(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Yue, Xianhu; Gu, Mingyong; Jia, TanghongBackground. Since the abnormal expression of miR-125b in spinal cord injury (SCI) and the regulatory effect of miR-125b on the MAPK pathway have been expounded, we attempt to investigate whether miR-125b exerts a regulatory effect on SCI by modulating the MAPK pathway. Method. A SCI rat model was established. The rats were treated with miR-125b antagomir or agomir, and their motor function affected by miR-125b was further detected by Basso-Beattie-Bresnahan (BBB) scoring. The histopathological changes and neuronal loss in the spinal cord were evaluated using hematoxylin-eosin and Nissl staining. Microglia-conditioned medium (MCM) was prepared and further used to treat the astrocytes, the activation of which was evaluated via immunofluorescence staining. The expressions of miR-125b, inflammation-related factors (IL-6, IL-1β, TNF-α, and IL-10), and MAPK pathway-related proteins (p38, ERK1/2, and JNK1/2 as well as their phosphorylated (p) forms) in the spinal cord, serum, and MCM-treated astrocytes of rats were determined by reversetranscription quantitative polymerase chain reaction (RTqPCR), enzyme-linked immunosorbent assay, and Western blot. Result. MiR-125b was lowly expressed in SCImodeled rats. MiR-125b downregulation aggravated the impaired motor function, the disorder within the tissue, astrocyte activation, and neuron loss in the spinal cord tissues of SCI-modeled rats, while miR-125b upregulation did oppositely. MiR-125b downregulation enhanced the levels of IL-6, IL-1β, TNF-α, p38, p-p38, p-ERK1/2, and p-JNK1/2, whilst reducing that of IL-10. Contrarily, miR-125b upregulation exerted the opposite effects in SCI-modeled rats and MCM-treated astrocytes. Conclusion. Up-regulation of miR-125b mitigates inflammation, astrocyte activation, and dysfunction in SCI by inactivating the MAPK pathway.
- PublicationOpen AccessSpatial transcriptomic analysis of tumour with high and low CAIX expression in TNBC tissue samples using GeoMx™ RNA assay(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Shamis, Suad A.K.; Savioli, Francesca; Ammar, Aula; Al Badran, Sara S.F.; Hatthakarnkul, Phimmada; Leslie, Holly; Mallon, Elizabeth E.A.; Jamieson, Nigel B.; McMillan, Donald C; Edwards, JoannePurpose. Prognostic significance and gene signatures associated with carbonic anhydrase IX (CAIX) was investigated in triple negative breast cancer (TNBC) patients. Methods. Immunohistochemistry (IHC) for CAIX was performed in tissue microarrays (TMAs) of 136 TNBC patients. In a subset of 52 patients Digital Spatial Profiler (DSP) was performed in tumour (pancytokeratin+) and stroma (pan-cytokeratin-). Differentially expressed genes (DEGs) with P<0.05 and log2 fold change (FC)>(±0.25 and ±0.3, for tumour and stromal compartment, respectively) were identified. Four genes were validated at the protein level. Result. Cytoplasmic CAIX expression was independently associated with poor recurrence free survival in TNBC patients [hazard ratio (HR)=6.59, 95% confidence interval (CI): 1.47-29.58, P=0.014]. DEG analysis identified 4 up-regulated genes (CD68, HIF1A, pan-melanocyte, and VSIR) in the tumour region and 9 down-regulated genes in the stromal region (CD86, CD3E, MS4A1, BCL2, CCL5, NKG7, PTPRC, CD27, and FAS) when low versus high CAIX expression was explored. Employing IHC, high CD68 and HIF-1α was associated with poorer prognosis and high BCL2 and CD3 was associated with good prognosis. Conclusions. DSP technology identified DEGs in TNBC. Selected genes validated by IHC showed involvement of CD3 and BCL2 expression within stroma and HIF-1α, and CD68 expression within tumour. However, further functional analysis is warranted
- PublicationOpen AccessExtracellular vesicles in acute respiratory distress syndrome: Understanding protective and harmful signaling for the development of new therapeutics(Universidad de Murcia. Departamento de Biología Celular e Histología, 2024) Bavuso, Matthew; Miller, Noel; Sill, Joshua M.; Dobrian, Anca; Colunga Biancatelli, Ruben M.L.Acute respiratory distress syndrome (ARDS) is a severe respiratory condition characterized by increased lung permeability, hyper-inflammatory state, and fluid leak into the alveolar spaces. ARDS is a heterogeneous disease, with multiple direct and indirect causes that result in a mortality of up to 40%. Due to the ongoing Covid-19 pandemic, its incidence has increased up to ten-fold. Extracellular vesicles (EVs) are small liposome-like particles that mediate intercellular communication and play a major role in ARDS pathophysiology. Indeed, they participate in endothelial barrier dysfunction and permeability, neutrophil, and macrophage activation, and also in the development of a hypercoagulable state. A more thorough understanding of the variegated and cell-specific functions of EVs may lead to the development of safe and effective therapeutics. In this review, we have collected evidence of EVs role in ARDS, revise the main mechanisms of production and internalization and summarize the current therapeutical approaches that have shown the ability to modulate EV signaling.
- PublicationOpen AccessPredictive markers for pathological complete response (pCR) after neo-adjuvant chemotherapy in HER2-positive breast carcinoma(Universidad de Murcia. Departamento de Biología Celular e Histología, 2024) Van Bockstal, Mieke R.; Dano, Hélène; Benhaddi, Naima; Dubois, Dominique; Vanderveken, Jonathan; Van Marcke, Cédric; Vandermeulen, Ad; Duhoux, Francois P.; Vernaeve, Hilde; Berlière, Martine; Galant, Christine
- PublicationOpen AccessAbnormal p16 expression and prognostic significance in esophageal squamous cell carcinoma(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Jiang, Dongxian; Zhang, Xue; Chen, Weijie; Peng, Rui; Zhang, Xiaolei; Gao, Feng; Huang, Yufeng; Gu, Wenyi; Hou, YingyongBackground. The purpose of this study was to analyze p16 expression status and evaluate whether abnormal p16 expression was associated with prognosis in a large-scale esophageal squamous cell carcinoma (ESCC) cohort of patients. Methods. We retrospectively evaluated p16 expression status of 525 ESCC samples using immunohistochemistry. Associations between abnormal p16 expression and survival were analyzed. Results. P16 negative, focal expression and overexpression were found in 87.6%, 6.9% and 5.5% of ESCC patients. No significant association was observed between abnormal p16 expression and age, sex, tumor site and location, differentiation, vessel and nerve invasion, T stage and lymph node metastasis. In all patients, the survival of p16 focal expression group tended to be better compared with negative group (disease free survival/DFS P=0.040 and overall survival/OS P=0.052) and overexpression group (DFS P=0.201 and OS P=0.258), and there was no survival difference between negative group and overexpression group. The multivariate analysis for OS and DFS found that only clinical stage was a significantly independent prognostic factor (P<0.001). When patients were divided into I-II stage (n=290) and III-IVa stage (n=235), the survival of focal expression group was better compared with negative group (DFS P=0.015 and OS P=0.019), and tended to be better compared with overexpression group (DFS P=0.405 and OS P=0.432) in I-II stage ESCC, which was not found in III-IVa stage ESCC. Conclusion. P16 overexpression or negative expression tend to be associated with unfavorable outcomes, especially in I-II stage ESCC. Our study will help to identify a subgroup of ESCC patients with excellent prognosis after surgical therapy
- PublicationOpen AccessResearch progress on the correlation between platelet aggregation and tumor progression(Universidad de Murcia. Departamento de Biología Celular e Histología, 2024) Chen, Yuyu; Yuan, Jialong; Tang, Faqing; Liu, Qinglin; Huang, Hongjun; Liu, Huan; Liu, HaoPlatelets are generally considered as the main functional unit of the coagulation system. However, more and more studies have confirmed that platelets also have an important relationship with tumor progression. Tumor cells can utilize platelets to promote their own infiltration and hematogenous metastasis, and platelets are activated and aggregated in this process. Therefore, platelet aggregation may be a concomitant marker of tumor progression. This is of great significance for predicting tumor metastasis before timely treatments.