Expression Of Sox2 Research Articles

Abstract A112: Mechanisms underlying platinum-induced ovarian cancer stem cell plasticity

Abstract Most women diagnosed with late-stage high grade serous ovarian cancer (HGSOC) develop recurrent, platinum-resistant tumors. Ovarian cancer stem cells (OCSCs) are hypothesized to contribute to the emergence of these resistant tumors. CSCs have been postulated to reside in a plastic state, which can allow for the conversion of non-CSC to CSC. This process of dedifferentiation continues during tumor development and chemotherapeutic agents like platinum can exaggerate CSC plasticity. We have previously demonstrated that acute platinum treatment enriched for OCSCs. However, whether platinum transforms non-OCSCs into OCSCs to contribute to this subpopulation of cells remains unclear, and the underlying mechanism remains incompletely understood. To examine OCSC plasticity, aldehyde dehydrogenase (ALDH; functional marker) and fluorescence activated cell sorting were used to isolate OCSCs (ALDH+) and non-OCSCs (ALDH-) from HGSOC cell lines, OVCAR5 and OVCAR3. To determine the stability of the non-OCSC phenotype, ALDH- cells were cultured for 3 and 5 days and ALDH activity was measured using flow cytometry. At both timepoints examined, ALDH- cells remained ALDH-, with approximately less than 1% being ALDH+ (p<0.05). To determine if platinum can induce conversion of non-OCSC to OCSC, ALDH- cells were treated with cisplatin (12µM or 15 µM for 16h), and the percent of ALDH+ cells was measured using flow cytometry. Treatment of ALDH- cells with cisplatin resulted in conversion of approximately 4% and 10% of ALDH- cells into ALDH+ cells (p<0.05) in OVCAR5 and OVCAR3 cells, respectively. Furthermore, both cell lines displayed increased expression (p<0.05) of stemness genes BMI1, NANOG, OCT4, and SOX2 and spheroid formation was observed in converted OCSCs compared to parental ALDH+ and whole cell populations treated with cisplatin, suggesting that platinum induced the observed differences in the stemness phenotype. With the goal of targeting key genes and pathways to inhibit platinum-induced OCSC conversion, RNA-sequencing was performed using converted OCSCs. Activated pathways in converted OCSCs were identified, including protein kinase signaling, inhibition of matrix metalloproteases, SPINK1 pathway, and VDR/RXR activation. We are further investigating if these pathways contribute to the mechanism involved in OCSC plasticity and whether targeting these pathways can reduce the OCSC population, which is critical to blocking the persistence of OCSCs and ultimately reduce mortality in patients. Citation Format: Tara X. Metcalfe, Shu Zhang, Christiane Hassel, Heather M. O'Hagan, Kenneth P. Nephew. Mechanisms underlying platinum-induced ovarian cancer stem cell plasticity [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A112.

A Comprehensive Understanding of Post-Translational Modification of Sox2 via Acetylation and O-GlcNAcylation in Colorectal Cancer.

Aberrant expression of the pluripotency-associated transcription factor Sox2 is associated with poor prognosis in colorectal cancer (CRC). We investigated the regulatory roles of major post-translational modifications in Sox2 using two CRC cell lines, SW480 and SW620, derived from the same patient but with low and high Sox2 expression, respectively. Acetylation of K75 in the Sox2 nuclear export signal was relatively increased in SW480 cells and promotes Sox2 nucleocytoplasmic shuttling and proteasomal degradation of Sox2. LC-MS-based proteomics analysis identified HDAC4 and p300 as binding partners involved in the acetylation-mediated control of Sox2 expression in the nucleus. Sox2 K75 acetylation is mediated by the acetyltransferase activity of CBP/p300 and ACSS3. In SW620 cells, HDAC4 deacetylates K75 and is regulated by miR29a. O-GlcNAcylation on S246, in addition to K75 acetylation, also regulates Sox2 stability. These findings provide insights into the regulation of Sox2 through multiple post-translational modifications and pathways in CRC.

A new animal model of lumbar disc degeneration in rabbits

BACKGROUND CONTEXTThere are many models of lumbar disc degeneration, but mechanical stress-induced lumbar disc degeneration is rare. Here we propose a mechanical stress-induced lumbar disc degeneration model to better understand the molecular mechanism of lumbar disc degeneration under stress stimulation. PURPOSETo design a new model of lumbar disc degeneration under mechanical stress. STUDY DESIGNThe anatomic approach of the oblique lateral approach to lumbar fusion surgery was used to design a longitudinal compression device across the vertebral body of the rabbit to impose longitudinal load on the lumbar disc. METHODSNew Zealand white rabbits (n=30) were used. Screws were used to cross the rabbits’ lumbar vertebral bodies, and both sides of the screws were pressurized. Continuous compression was then performed for 28 days. Adjacent unpressurized lumbar discs serve as controls for pressurized lumbar discs. At 28 days after surgery, micro-computed tomography (CT) and magnetic resonance imaging (MRI) were performed on the rabbits’ lumbar discs. After the imaging examination, lumbar disc samples were removed, Safranin-O fast green and immunofluorescence was performed to detect the expression level of intervertebral disc degeneration-related proteins. RESULTSThe CT results showed that the disc height did not decrease significantly after mechanical loading. The MRI results showed that the signals in the pressurized disc decreased 28 days after loading. The results of Safranin-O fast green showed that the cartilage component of the intervertebral disc after mechanical compression was significantly reduced. The immunofluorescence results showed that the expression of ADAMTS5 and MMP13 protein in the nucleus pulposus of the intervertebral disc after mechanical compression increased, while the expression of SOX9 decreased, and the difference was statistically significant. Aggrecan's protein expression decreased, but was not statistically significant. CONCLUSIONSThis study designed a reliable model of disc degeneration in rabbits. It is more likely to mimic disc compression in the human body. CLINICAL SIGNIFICANCEThis animal model can be used as a basic model to study the molecular physiological mechanisms of discogenic low back pain.

SOX2 and OCT4 mediate radiation and drug resistance in pancreatic tumor organoids

Pancreatic cancer has a five-year survival rate of only 10%, mostly due to late diagnosis and limited treatment options. In patients with unresectable disease, either FOLFIRINOX, a combination of 5-fluorouracil (5-FU), oxaliplatin and irinotecan, or gemcitabine plus nab-paclitaxel combined with radiation are frontline standard regimens. However, chemo-radiation therapy has shown limited success because patients develop resistance to chemotherapy and/or radiation. In this study, we evaluated the role of pancreatic cancer stem cells (CSC) using OCT4 and SOX2, CSC markers in mouse pancreatic tumor organoids. We treated pancreatic tumor organoids with 4 or 8 Gy of radiation, 10 μM of 5-FU (5-Fluorouracil), and 100 μM 3-Bromopyruvate (3BP), a promising anti-cancer drug, as a single treatment modalities, and in combination with RT. Our results showed significant upregulation of, OCT4, and SOX2 expression in pancreatic tumor organoids treated with 4 and 8 Gy of radiation, and downregulation following 5-FU treatment. The expression of CSC markers with increasing treatment dose exhibited elevated upregulation levels to radiation and downregulation to 5-FU chemotherapy drug. Conversely, when tumor organoids were treated with a combination of 5-FU and radiation, there was a significant inhibition in SOX2 and OCT4 expression, indicating CSC self-renewal inhibition. Noticeably, we also observed that human pancreatic tumor tissues exhibited heterogeneous and aberrant OCT4 and SOX2 expression as compared to normal pancreas, indicating their potential role in pancreatic cancer growth and therapy resistance. In addition, the combination of 5-FU and radiation treatment exhibited significant inhibition of the β-catenin pathway in pancreatic tumor organoids, resulting in sensitization to treatment and organoid death. In conclusion, our study emphasizes the crucial role of CSCs in therapeutic resistance in PC treatment. We recommend using tumor organoids as a model system to explore the impact of CSCs in PC and identify new therapeutic targets.

Expression of SOX4 Significantly Predicts the Risk of Lymph Node Metastasis for Patients With Early-Stage Esophageal Squamous Cell Carcinoma

Esophageal squamous cell carcinoma stands as a notably aggressive malignancy within the digestive system. In cases of early esophageal cancer without lymph node metastasis, endoscopic surgical resection offers a viable alternative, often resulting in improved patient quality of life. However, the paucity of methods to preoperatively ascertain lymph node involvement complicates surgical planning. SOX4 gene was previously found to be highly associated with invasive metastasis in our work through single-cell RNA sequencing on 5 paired tumor/peritumor tissues. This research included the collection of 124 tissue samples from 106 patients (106 tumor and 18 lymph node specimens). Samples were methodically arranged into a tissue microarray and treated with immunohistochemical staining. Statistical analysis was conducted to assess the relationship between them. In the univariate analysis, 3 factors were identified as statistically significant in relation to lymph node metastasis: T category (P = .014), vascular invasion (P < .001), and SOX4 intensity (P = .001). Additionally, when evaluating SOX4 intensity alongside other clinical indicators, SOX4 was shown to independently influence lymph node metastasis. Further, the multivariate analysis revealed that vascular invasion (P < .001) and SOX4 intensity (P = .003) were significantly associated with lymph node metastasis, exhibiting hazard ratios of 10.174 and 7.142, respectively. The results of our study indicate that both SOX4 expression and vascular invasion serve as predictors of lymph node metastasis in patients diagnosed with category T1 esophageal squamous cell carcinoma, underscoring the potential utility of SOX4 in prognostic evaluations.

Role of sodium/iodide symporter overexpression in inhibiting thyroid cancer cell invasion and stem cell maintenance by inhibiting the β-catenin/LEF-1 pathway

BackgroundIn thyroid cancers, a reduction in the expression of the sodium/iodide symporter (NIS) is observed concomitant with a diminution in cancer cell differentiation. The β-catenin/LEF-1 pathway emerges as a crucial regulatory pathway influencing the functional expression of NIS in human thyroid cancer cells. Further research is required to comprehensively elucidate the role of NIS overexpression in impeding the progression of thyroid cancer cells. MethodsHuman papillary thyroid carcinoma (PTC) cell lines, specifically PTC-1 and KTC-1, were subjected to Scratch and Transwell assays, colony formation, and tumor sphere formation tests to investigate invasion and migration, focusing on the impact of NIS overexpression. The assessment involved the use of western blot to analyze the expression levels of β-catenin, NIS, CD133, SRY-related HMG box2 (Sox2), lymphoid enhancer-binding factor 1 (LEF-1), NANOG, octamer-binding transcription factor 4 (Oct4), aldehyde dehydrogenase 1 family, member A1 (ALDH1A1), and epithelial cellular adhesion molecule (EpCAM). Statistical analysis was conducted using SPSS version 20.0, and the graphs were developed using GraphPad Prism 7 (GraphPad Software, Inc.). ResultsOur observations revealed that Nthy-ori-3-1 cell lines exhibited notably higher average expression levels of NIS, yet significantly lower levels of LEF-1 and β-catenin compared to PTC-1 and KTC-1 cell lines. Furthermore, the overexpression of β-catenin resulted in reduced binding of LEF-1 to NIF promotion but concurrently increased the expression of NIS. The downregulation of NIS markedly enhanced the expression of ALDH1A1, CD133, OCT4, Nanog, SOX2, and EpCam—all of which are targets within the Wnt/β-catenin signaling pathway. Conversely, the upregulation of NIS suppressed the expression of these proteins. Moreover, cells treated with β-catenin activators demonstrated an increased capability to form more spheroids and displayed heightened aggressiveness. Conversely, the NIS overexpression (OE) group exhibited suppressed abilities in invasion and colony formation. ConclusionThyroid cancer cells exhibit diminished expression of NIS, and the invasion and maintenance of stem cells in thyroid cancer cells were hindered by NIS OE through the inhibition of the β-catenin/LEF-1 pathway. Further research is warranted to comprehensively assess this outcome, which holds promise as a potential targeted treatment for thyroid cancer.

MiR-29a-KLF4 signaling inhibits breast tumor initiation by regulating cancer stem cells

Cancer stem cells (CSCs) are known for their potent ability to drive tumor initiation and recurrence, yet the molecular mechanisms regulating CSCs are still unclear. Our study found a positive correlation between increased levels of miR-29a and better survival rates in early-stage breast cancer patients, but a negative correlation in late-stage patients, suggesting a dual function of miR-29a in regulating breast cancer. Furthermore, miR-29a showed significant downregulation in the ALDH+ breast cancer stem cell population compared to non-stem cancer cells. Overexpression of miR-29a in human breast cancer cells reduced the proportion of CSCs, suppressed their ability to form mammospheres, and inhibited the expression of stemness genes SOX2, KLF4, and hTERT in vitro. Conversely, knockdown of miR-29a in breast cancer cells showed opposite effects. Tumor xenograft experiments revealed that miR-29a overexpression significantly inhibited tumorigenesis initiated by MDA-MB-231 cell transplantation in nude mice. We further demonstrated that Krüppel-like factor 4 (KLF4), a key gene that regulates cell stemness, was a direct target of miR-29a in breast cancer cells. miR-29a suppressed the expression of KLF4 at both mRNA and protein levels. Reintroduction of KLF4 into breast cancer cells rescued the miR-29a-induced CSC suppression phenotype. In summary, our study is the first to demonstrate that miR-29a-KLF4 signaling inhibits breast tumor initiation by regulating CSCs, which provides novel therapeutic targets for preventing breast tumor initiation.

Pcgf5: An important regulatory factor in early embryonic neural induction

Polycomb group RING finger (PCGF) proteins, a crucial subunits of the Polycomb complex, plays an important role in regulating gene expression, embryonic development, and cell fate determination. In our research, we investigated Pcgf5, one of the six PCGF homologs, and its impact on the differentiation of P19 cells into neural stem cells. Our findings revealed that knockdown of Pcgf5 resulted in a significant decrease in the expression levels of the neuronal markers Sox2, Zfp521, and Pax6, while the expression levels of the pluripotent markers Oct4 and Nanog increased. Conversely, Pcgf5 overexpression upregulated the expression of Sox2 and Pax6, while downregulating the expression of Oct4 and Nanog. Additionally, our analysis revealed that Pcgf5 suppresses Wnt3 expression via the activation of Notch1/Hes1, and ultimately governs the differentiation fate of neural stem cells. To further validate our findings, we conducted in vivo experiments in zebrafish. We found that knockdown of pcgf5a using morpholino resulted in the downregulated expression of neurodevelopmental genes such as sox2, sox3, and foxg1 in zebrafish embryos. Consequently, these changes led to neurodevelopmental defects. In conclusion, our study highlights the important role of Pcgf5 in neural induction and the determination of neural cell fate.

Endothelial cells drive organ fibrosis in mice by inducing expression of the transcription factor SOX9.

Fibrosis is a hallmark of chronic disease. Although fibroblasts are involved, it is unclear to what extent endothelial cells also might contribute. We detected increased expression of the transcription factor Sox9 in endothelial cells in several different mouse fibrosis models. These models included systolic heart failure induced by pressure overload, diastolic heart failure induced by high-fat diet and nitric oxide synthase inhibition, pulmonary fibrosis induced by bleomycin treatment, and liver fibrosis due to a choline-deficient diet. We also observed up-regulation of endothelial SOX9 in cardiac tissue from patients with heart failure. To test whether SOX9 induction was sufficient to cause disease, we generated mice with endothelial cell-specific overexpression of Sox9, which promoted fibrosis in multiple organs and resulted in signs of heart failure. Endothelial Sox9 deletion prevented fibrosis and organ dysfunction in the two mouse models of heart failure as well as in the lung and liver fibrosis mouse models. Bulk and single-cell RNA sequencing of mouse endothelial cells across multiple vascular beds revealed that SOX9 induced extracellular matrix, growth factor, and inflammatory gene expression, leading to matrix deposition by endothelial cells. Moreover, mouse endothelial cells activated neighboring fibroblasts that then migrated and deposited matrix in response to SOX9, a process partly mediated by the secreted growth factor CCN2, a direct SOX9 target; endothelial cell-specific Sox9 deletion reversed these changes. These findings suggest a role for endothelial SOX9 as a fibrosis-promoting factor in different mouse organs during disease and imply that endothelial cells are an important regulator of fibrosis.

Effects of self-assembled type II collagen fibrils on the morphology and growth of pre-chondrogenic ATDC5 cells

ObjectiveAlthough type II collagen could have marked potential for developing cartilage tissue engineering (CTE) scaffolds, its erratic supply and viscous nature have limited these studies, and there are no studies on the use of marine-derived type II collagen fibrils for CTE scaffold materials. In this study, we aimed to generate a fibril-based, thin-layered scaffold from marine-derived type II collagen and investigate its chondrogenic potential. MethodsTime-lapse observations revealed the cell adhesion process. The Cell Counting Kit-8 (CCK-8) assay, light microscopy, and scanning electron microscopy were performed to detect proliferation and filopodium morphology. Alcian blue staining was used to show the deposition of extracellular secretions, and qRT-PCR was performed to reveal the expression levels of chondrogenesis-related genes. ResultsThe cell adhesion speed was similar in both fibril-coated and control molecule-coated groups, but the cellular morphology, proliferation, and chondrogenesis activity differed. On fibrils, more elongated finer filopodia showed inter-cell communications, whereas the slower proliferation suggested an altered cell cycle. Extracellular secretions occurred before day 14 and continued until day 28 on fibrils, and on fibrils, the expression of the chondrogenesis-related genes Sox9 (p ​< ​0.001), Col10a1 (p ​< ​0.001), Acan (p ​< ​0.001), and Col2a1 (p ​= ​0.0049) was significantly upregulated on day 21. ConclusionMarine-derived type II collagen was, for the first time, fabricated into a fibril state. It showed rapid cellular affinity and induced chondrogenesis with extracellular secretions. We presented a new model for studying chondrogenesis in vitro and a potential alternative material for cell-laden CTE research.

CHD7 and SOX2 act in a common gene regulatory network during mammalian semicircular canal and cochlear development

Inner ear morphogenesis requires tightly regulated epigenetic and transcriptional control of gene expression. CHD7, an ATP-dependent chromodomain helicase DNA-binding protein, and SOX2, an SRY-related HMG box pioneer transcription factor, are known to contribute to vestibular and auditory system development, but their genetic interactions in the ear have not been explored. Here, we analyzed inner ear development and the transcriptional regulatory landscapes in mice with variable dosages of Chd7 and/or Sox2. We show that combined haploinsufficiency for Chd7 and Sox2 results in reduced otic cell proliferation, severe malformations of semicircular canals, and shortened cochleae with ectopic hair cells. Examination of mice with conditional, inducible Chd7 loss by Sox2CreER reveals a critical period (~E9.5) of susceptibility in the inner ear to combined Chd7 and Sox2 loss. Data from genome-wide RNA-sequencing and CUT&Tag studies in the otocyst show that CHD7 regulates Sox2 expression and acts early in a gene regulatory network to control expression of key otic patterning genes, including Pax2 and Otx2. CHD7 and SOX2 directly bind independently and cooperatively at transcription start sites and enhancers to regulate otic progenitor cell gene expression. Together, our findings reveal essential roles for Chd7 and Sox2 in early inner ear development and may be applicable for syndromic and other forms of hearing or balance disorders.

Cancer stem cell biomarkers SOX2 and Oct4 in cervical cancer patients undergoing chemoradiotherapy.

Cancer stem cell biomarkers SRY (sex-determining region Y)-box 2 (SOX2) and octamer-binding transcription factor 4 (Oct4) account for radioresistance in cervical squamous cell cancers (CSCCs). Their clinical implications are limited and contradictory. In this prospective cohort study, we recruited patients with FIGO IB2-IVA CSCC treated with primary chemoradiotherapy on regular follow-up. Tissue biopsy specimens were evaluated for SOX2 and Oct4 expression by immunohistochemistry, quantified by a product of proportion and intensity scores. A total of 59 patients were included. Most had a moderately differentiated (81%), keratinizing (59%) CSCC, and ≥FIGO stage IIB disease (95%). SOX2 expression (high:low 21:38 patients) and Oct4 expression (high:low 4:55 patients) had a significant interrelation (p=0.005, odds ratio (95% CI) - 1.23 (1.004-1.520)). At a median follow-up of 36 months, the 3-year overall survival (OS) was 60% and 53% for low and high SOX2 expression (p=0.856), and 54% and 100% for low and high Oct4 expression (p=0.114). The 3-year disease-frese survival (DFS) was 65% and 50% in the low and high SOX2 expression (p=0.259), and 59% and 75% for low and high Oct4 expression (p=0.598). SOX2 expression was the only variable significantly associated with a lower OS and DFS on regression analysis. Our study demonstrated a trend toward improved OS and DFS with low SOX2 and high Oct4 expression in CSCC patients undergoing chemoradiotherapy.

Preliminary evaluation of fish cartilage as a promising biomaterial in cartilage tissue engineering

Fish cartilage is known as a valuable source of natural biomaterials due to its unique composition and properties. It contains a variety of bioactive components that contribute to its potential applications in different domains such as tissue engineering. The present work aimed to consider the properties of backbone cartilage from fish with a cartilaginous skeleton, including elasmobranch (reticulate whipray: Himantura uarnak and milk shark: Rhizoprionodon acutus) and sturgeon (beluga: Huso huso). The histomorphometric findings showed that the number of chondrocytes was significantly higher in reticulate whipray and milk shark compared to beluga (p < 0.05). The highest GAGs content was recorded in reticulate whipray cartilage compared to the other two species (p < 0.05). The cartilage from reticulate whipray and beluga showed higher collagen content than milk shark cartilage (p < 0.05), and the immunohistochemical assay for type II collagen (Col II) showed higher amounts of this component in reticulate whipray compared to the other two species. Young’s modulus of the cartilage from reticulate whipray was significantly higher than that of milk shark and beluga (p < 0.05), while no significant difference was recorded between Young’s modulus of the cartilage from milk shark and beluga. The gene expression of ACAN, Col II, and Sox9 showed that the cartilage-ECM from three species was able to induce chondrocyte differentiation from human adipose tissue-derived stem cells (hASCs). From these results, it can be concluded that the cartilage from three species, especially reticulate whipray, enjoys the appropriate biological properties and provides a basis for promoting its applications in the field of cartilage tissue engineering.

Kurarinone targets JAK2-STAT3 signaling in colon cancer-stem-like cells.

Natural compounds are known to regulate stemness/self-renewal properties in colon cancer cells at molecular level. In the present study, we first time studied the colon cancer stem-like cells targeting potential of Kurarinone (KU) and explored the underlying mechanism. Cytotoxic potential of KU was checked in colon cancer cells. Colonosphere formation assay was performed to check the spheroid formation reduction potential of KU in HCT-116 cells by using phase-contrast microscopy. Stemness/self-renewal marker expression was studied at mRNA and protein levels in colonosphere. The qRT-PCR, western blot analysis, and flow cytometer techniques were used to assess the effect of KU treatment on cell cycle progression and apoptosis induction in colon cancer cells and colonosphere. Further, effect of KU treatment on pSTAT3 status and its nuclear translocation was also studied. KU treatment significantly decreased HCT-116 cell proliferation and reduced sphere formation potential at IC30 (8.71 µM) and IC50 (20.34 µM) concentrations compared to respective vehicle-treated groups, respectively. KU exposure significantly reduced the expression of CD44, c-Myc, Bmi-1, and Sox2 stemness/self-renewal markers in colonosphere in a dose-dependent manner. KU treatment inhibits JAK2-STAT3 signaling pathway by reducing pSTAT3 levels and its nuclear translocation in HCT-116 cells and colonosphere at IC50 concentration. KU treatment significantly decreased the expression of CCND1 and CDK4 cell cycle-specific markers and arrested the HCT-116 cells and colonosphere in G1-phase. Further, KU treatment increased Bax/Bcl-2 ratio, apoptotic cell population, cleaved caspase 3, and PARP-1 in HCT-116 cells and colonosphere. In conclusion, KU treatment decreases stemness/self-renewal, induces cell cycle arrest and apoptosis in HCT-116 colonosphere by down-regulating CD44-JAK2-STAT3 axis. Thus, targeting stemness/self-renewal and other cancer hallmark(s) by KU through CD44/JAK2/STAT3 signaling pathway might be a novel strategy to target colon cancer stem-like cells.

MFAP5 Strengthened the Stem Cell Features of Non-small Cell Lung Cancer Cells by Regulating the FBW/Sox9 Axis.

Non-small cell lung cancer (NSCLC) is a type of malignant tumor with high morbidity as well as mortality. The process of lung cancer may be driven by cancer stem cells. It was known that MFAP5 enhanced the occurrence of diverse types of cancer. Also, MFAP5 has the potential to induce the degradation of FBW7 which is a tumor suppressor. Lower levels of FBW7 enhance the stability of Sox9, which is the cancer stem cell-related protein. However, whether the MFAP5 can modulate the stem cell features of NSCLC cells by modulating the FBW7/Sox9 axis is unclear. Therefore, this study aimed to explore the role of MFAP5/FBW7/Sox9 axis on the stem cell features of NSCLC cells and develop a new treatment of this carcinoma. In this study, we explored the effects of MFAP5 on the stem cell features of NSCLC cells for the first time. We established MFAP5 overexpression and knockdown NSCLC cells. Clone formation assays and cell sphere culture assays were conducted for the exploration of the growth and stem cell features of these cells. Western blotting was applied for the detection of Sox9 and FBW7 expression in these cells. CHX was applied for the treatment of these cells for the detection of degradation of Sox9. Finally, we overexpressed the Sox9 in MFAP5 knockdown NSCLC cells. MFAP5 promoted the growth and stem cell features of these cells. Knockdown of MFAP5 induced higher levels of FBW7 while restricting the expression of Sox9. Knockdown of MFAP5 aggravated the degradation of Sox9. Overexpression of Sox9 abrogated the efficacy of MFAP5 inhibition on the growth as well as stem cell features of these cells. The results of this study clarified the role of MFAP5/FBW7/Sox9 axis on the development of non-small cell lung cancer cells, providing the potential therapeutic target for the clinical treatment of NSCLC. MFAP5 maintained the stem cell features of non-small cell lung cancer cells by modulating FBW7/Sox9 axis.

Logic "AND Gate Circuit"-Based Gasdermin Protein Expressing Nanoplatform Induces Tumor-Specific Pyroptosis to Enhance Cancer Immunotherapy.

Pyroptosis mediated by gasdermin protein has shown great potential in cancer immunotherapies. However, the low expression of gasdermin proteins and the systemic toxicity of nonspecific pyroptosis limit its clinical application. Here, we designed a synthetic biology strategy to construct a tumor-specific pyroptosis-inducing nanoplatform M-CNP/Mn@pPHS, in which a pyroptosis-inducing plasmid (pPHS) was loaded onto a manganese (Mn)-doped calcium carbonate nanoparticle and wrapped in a tumor-derived cell membrane. M-CNP/Mn@pPHS showed an efficient tumor targeting ability. After its internalization by tumor cells, the degradation of M-CNP/Mn@pPHS in the acidic endosomal environment allowed the efficient endosomal escape of plasmid pPHS. To trigger tumor-specific pyroptosis, pPHS was designed according to the logic "AND gate circuit" strategy, with Hif-1α and Sox4 as two input signals and gasdermin D induced pyroptosis as output signal. Only in cells with high expression of Hif-1α and Sox4 simultaneously will the output signal gasdermin D be expressed. Since Hif-1α and Sox4 are both specifically expressed in tumor cells, M-CNP/Mn@pPHS induces the tumor-specific expression of gasdermin D and thus pyroptosis, triggering an efficient immune response with little systemic toxicity. The Mn2+ released from the nanoplatform further enhanced the antitumor immune response by stimulating the cGAS-STING pathway. Thus, M-CNP/Mn@pPHS efficiently inhibited tumor growth with 79.8% tumor regression in vivo. We demonstrate that this logic "AND gate circuit"-based gasdermin nanoplatform is a promising strategy for inducing tumor-specific pyroptosis with little systemic toxicity.

Assessment of SOX10 expression in 437 canine neoplasms of different embryologic origins.

Several members of the SRY-related HMG-box (SOX) protein family are implicated in tumorigenesis, metastasis, and regulation of the tumor microenvironment. SOX10, which is involved in neural crest cell migration and differentiation, has long been recognized a sensitive and specific immunohistochemical (IHC) marker in the diagnosis of melanoma in humans. However, expression of SOX10 in other tumor types has infrequently been evaluated in humans until recently and has not been thoroughly investigated in the dog. Our aim was to characterize the expression of SOX10 in canine neoplasms to objectively assess its value as a diagnostic IHC marker. Immunohistochemistry for SOX10 was performed on 437 archived, formalin-fixed paraffin-embedded tissues from representative canine neoplasms of ectodermal (15 tumor types), mesodermal (13 tumor types), endodermal (8 tumor types), and mixed/unknown (7 tumor types) embryologic origin. Oral and cutaneous tumors of melanocytic origin were used as positive controls. Intense SOX10 immunolabeling was observed in most tumors of ectodermal origin, including consistent expression in mammary carcinomas, and gliomas. Embryonal and hair follicle neoplasms inconsistently exhibited strong nuclear immunolabeling. Oral fibrosarcomas and undifferentiated oral sarcomas both inconsistently exhibited moderate to strong nuclear immunolabeling. Neoplasms of mesodermal and endodermal origin lacked immunolabeling. Salivary carcinomas, representing an unknown/mixed embryologic origin, were strongly labeled. SOX10 expression is not limited to melanomas, but is expressed by canine tumors of diverse tissues and embryologic derivation. Importantly, expression of SOX10 by a subset of oral sarcomas impairs its value as a marker for spindle cell oral melanomas.

The Role of P4HB and SOX4 in Prostatic Carcinoma and Their Clinical Significance.

Prostatic adenocarcinoma (PAC) is the second most prevalent cancer and the fifth leading cause of cancer death in men worldwide. Additionally, pathologists may face problems diagnosing it reliably and may need more than one marker. Thus, the search for new immunohistochemical biomarkers becomes mandatory. This study aims to investigate P4HB and SOX4 expression in prostatic carcinoma, their possible roles, and clinical significance. This retrospective study included fifty-six cases of PAC and an equal number of nodular prostatic hyperplasia (NPH) that were immunohistochemically stained by P4HB and SOX4. The results of expression were compared between PAC and NPH cases, followed by correlations with available clinicopathological parameters. There was a highly significant difference between PAC and NPH regarding P4HB and SOX4 expressions in favor of PAC (both P<0.001). ROC curve analysis of the diagnostic power of P4HB showed 79% sensitivity, 76% specificity, and an area under the ROC curve of 0.845, while SOX4 showed (89%, 100%, and 0.946, respectively). P4HB and SOX4 expression showed a direct correlation (P<0.001). Moreover, the H-score of SOX4 expression showed a significant inverse relation with ERG expression (P=0.047). There was a significant correlation between P4HB and SOX4 and Gleason score (P<0.001). Moreover, P4HB expression was significantly associated with lymphovascular invasion (P=0.013), while SOX4 expression showed a significant association with perineural invasion (P=0.05). SOX4 and P4HB seem to have diagnostic and prognostic value in PAC. While there was a direct correlation between SOX4 and P4HB, an inverse relationship between SOX4 and ERG was detected.

Skeletal growth is enhanced by a shared role for SOX8 and SOX9 in promoting reserve chondrocyte commitment to columnar proliferation

SOX8 was linked in a genome-wide association study to human height heritability, but roles in chondrocytes for this close relative of the master chondrogenic transcription factor SOX9 remain unknown. We undertook here to fill this knowledge gap. High-throughput assays demonstrate expression of human SOX8 and mouse Sox8 in growth plate cartilage. In situ assays show that Sox8 is expressed at a similar level as Sox9 in reserve and early columnar chondrocytes and turned off when Sox9 expression peaks in late columnar and prehypertrophic chondrocytes. Sox8-/- mice and Sox8fl/flPrx1Cre and Sox9fl/+Prx1Cre mice (inactivation in limb skeletal cells) have a normal or near normal skeletal size. In contrast, juvenile and adult Sox8fl/flSox9fl/+Prx1Cre compound mutants exhibit a 15 to 20% shortening of long bones. Their growth plate reserve chondrocytes progress slowly toward the columnar stage, as witnessed by a delay in down-regulating Pthlh expression, in packing in columns and in elevating their proliferation rate. SOX8 or SOX9 overexpression in chondrocytes reveals not only that SOX8 can promote growth plate cell proliferation and differentiation, even upon inactivation of endogenous Sox9, but also that it is more efficient than SOX9, possibly due to greater protein stability. Altogether, these findings uncover a major role for SOX8 and SOX9 in promoting skeletal growth by stimulating commitment of growth plate reserve chondrocytes to actively proliferating columnar cells. Further, by showing that SOX8 is more chondrogenic than SOX9, they suggest that SOX8 could be preferred over SOX9 in therapies to promote cartilage formation or regeneration in developmental and degenerative cartilage diseases.

Clinicopathological analysis of gonadal differentiation of sex development disorder

Objective: To investigate pathological features and differential diagnosis in the gonads with disorder of sex development. Methods: Thirty-six cases of clinically diagnosed hermaphroditism with gonadal biopsy in the Department of Pathology, the Seventh Medical Center of People's Liberation Army General Hospital from April 2007 to July 2021, were collected. All biopsy pathological sections were reviewed, and the gonadal cases with abnormal pathological morphology were screened out. The clinical and imaging data and karyotype of these cases were reviewed. Additional immunohistochemical staining was performed and relevant literature was reviewed. Results: Seven cases of ovotesticular disorder of sex development (OTDSD) were identified, which were characterized by the presence of testicular and ovarian differentiation in the same individual. All patients were under 15 years old and presented with abnormal appearance of external genitalia, and the ratio of male to female was 2∶5. Ultrasonography showed testicular structure in all female patients and cryptorchidism in all male patients. The most common karyotype was 46, XX. One case with undifferentiated gonadal tissue (UGT) and one case with streak gonads were screened out. UGT germ cells were neither in seminiferous tubules nor in follicles, but randomly distributed in an ovarial-type interstitial background, sometimes accompanied by immature sex cords. Streak gonads resembled UGT without germ cells. FOXL2 was positive in granulosa cells, but negative in Sertoli cells. SOX9 expression was opposite. OCT4 was weakly positively/negatively expressed in oocytes and positively expressed in the germ nuclei of UGT. Conclusions: Four differentiation patterns need to be identified in the gonadal biopsy: ovarian differentiation, testicular differentiation, undifferentiated gonadal tissue and streak gonad. The positive expression of SOX9 indicates testicular differentiation, while the positive expression of FOXL2 confirms ovarian differentiation, and the expression of both markers in the same tissue indicates ovotestis differentiation. It is very important to identify UGT, because that has a high probability of developing into gonadoblastoma in the future.