Evaluation of Paxillin Expression in Odontogenic Cysts and Tumors.

Statement of the Problem: Paxillin (PXN) is one of the proteins involved in cell adhesion. PXN and integrins constitute a key site for the focal adhesion between the cell and extracellular matrix. Several studies have shown that PXN is a factor in tumor formation, progression, invasion, and metastasis. Purpose: This study evaluated PXN expression in four types of odontogenic lesions with different aggressive behaviors. Materials and Method: In this retrospective cross-sectional study, PXN expression was immunohistochemically assessed in 68 paraffin-embedded tissue samples from patients with the confirmed diagnosis of four types of odontogenic lesions, including 14 dentigerous cysts (DC), 20 odontogenic keratocyst (OKC), 16 unicystic ameloblastoma, and 18 solid ameloblastoma. The PXN expression in these samples were scored based on the percentage and intensity of immunoreactivity, and compared among the groups by Chi-square test. Results: The PXN marker was detected in the cytoplasm of tumor cells (unicystic and solid ameloblastoma) and the epithelial layer of cysts (DC and OKC). The intensively stained marker of PXN was observed in 9 cases (64.3%) of the DC, 14 cases (70%) of OKC, 12 cases (75%) of unicystic ameloblastoma, and 13 cases (72.2%) of solid ameloblastoma. However, there was not statistical difference of PXN protein expression between DC and OKC (p Value = 0.51) and unicystic and solid ameloblastoma (p = 0.58), also the same was true for cysts and tumors (p = 0.37). Conclusion: The expression of PXN is not related to the biological behaviors of odontogenic lesions.

Plexin D1 negatively regulates macrophage-derived foam cell migration via the focal adhesion kinase/Paxillin pathway.

BACKGROUND: Macrophage-derived foam cell formation is a hallmark of atherosclerosis and is retained during plaque formation. Strategies to inhibit the accumulation of these cells hold promise as viable options for treating atherosclerosis. Plexin D1 (PLXND1), a member of the Plexin family, has elevated expression in atherosclerotic plaques and correlates with cell migration; however, its role in macrophages remains unclear. We hypothesize that the guidance receptor PLXND1 negatively regulating macrophage mobility to promote the progression of atherosclerosis. METHODS: We utilized a mouse model of atherosclerosis based on a high-fat diet and an ox-LDL- induced foam cell model to assess PLXND1 levels and their impact on cell migration. Through western blotting, Transwell assays, and immunofluorescence staining, we explored the potential mechanism by which PLXND1 mediates foam cell motility in atherosclerosis. RESULTS: Our study identifies a critical role for PLXND1 in atherosclerosis plaques and in a low-migration capacity foam cell model induced by ox-LDL. In the aortic sinus plaques of ApoE-/- mice, immunofluorescence staining revealed significant upregulation of PLXND1 and Sema3E, with colocalization in macrophages. In macrophages treated with ox-LDL, increased expression of PLXND1 led to reduced pseudopodia formation and decreased migratory capacity. PLXND1 is involved in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK. Additionally, FAK inhibitors counteract the ox-LDL-induced migration suppression by modulating the phosphorylation states of FAK, Paxillin and their downstream effectors CDC42 and PAK. CONCLUSION: Our findings indicate that PLXND1 plays a role in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK to promoting atherosclerosis.

Phytoconstituent-derived zingerone nanoparticles disrupt the cell adhesion mechanism and suppress cell motility in melanoma B16F10 cells.

Combining phytochemicals and nanotechnology to improve the unfavorable innate properties of phytochemicals and develop them into potent nanomedicines to enhance antitumor efficacy has become a novel strategy for cancer chemoprevention. Melanoma is the most aggressive, metastatic, and deadly disease of the primary cutaneous neoplasms. In this study, we fabricated phytoconstituent-derived zingerone nanoparticles (NPs) and validated their effects on cell adhesion and motility in melanoma B16F10 cells. Our data indicated that zingerone NPs significantly induced cytotoxicity and anti-colony formation and inhibited cell migration and invasion. Moreover, zingerone NPs dramatically interfered with the cytoskeletal reorganization and markedly delayed the period of cell adhesion. Our results also revealed that zingerone NPs-mediated downregulation of MMPs (matrix metalloproteinases) activity is associated with inhibiting cell adhesion and motility. We further evaluated the effects of zingerone NPs on Src/FAK /Paxillin signaling, our data showed that zingerone NPs significantly inhibited the protein activities of Src, FAK, and Paxillin, indicating that they play important roles in zingerone NP-mediated anti-motility and anti-invasion in melanoma cells. Accordingly, the phytoconstituent-zingerone NPs can strengthen the inhibition of tumor growth, invasion, and metastasis in malignant melanoma. Altogether, these multi-pharmacological benefits of zingerone NPs will effectively achieve the purpose of melanoma prevention and invasion inhibition.

LncRNA MALAT1 facilitates BM-MSCs differentiation into endothelial cells and ameliorates erectile dysfunction via the miR-206/CDC42/PAK1/paxillin signalling axis.

BACKGROUND: Erectile dysfunction (ED) is a common male sexual dysfunction, with an increasing incidence, and the current treatment is often ineffective. METHODS: Vascular endothelial growth factor (VEGFA) was used to treat bone marrow-derived mesenchymal stem cells (BM-MSCs), and their cell migration rates were determined by Transwell assays. The expression of the von Willebrand Factor (vWF)VE-cadherin, and endothelial nitric oxide synthase(eNOS) endothelial markers was determined by qRT‒PCR and Western blot analyses. The MALAT1-induced differentiation of BM-MCs to ECs via the CDC42/PAK1/paxillin pathway was explored by transfecting VEGFA-induced BM-MSC with si-MALAT1 and overexpressing CDC42 and PAK1. The binding capacity between CDC42, PAK1, and paxillin in VEGFA-treated and non-VEGFA-treated BM-MSCs was examined by protein immunoprecipitation. MiR-206 was overexpressed in VEGFA-induced BM-MSC, and the binding sites of MALAT1, miR-206, and CDC42 were identified using a luciferase assay. Sixty male Sprague‒Dawley rats were divided into six groups (n = 10/group). DMED modelling was demonstrated by APO experiments and was assessed by measuring blood glucose levels. Erectile function was assessed by measuring the intracavernosa pressure (ICP) and mean arterial pressure (MAP). Penile erectile tissue was analysed by qRT‒PCR, Western blot analysis, and immunohistochemical staining. RESULTS: MALAT1 under VEGFA treatment conditions regulates the differentiation of BM-MSCs into ECs by modulating the CDC42/PAK1/paxillin axis. In vitro experiments demonstrated that interference with CDC42 and MALAT1 expression inhibited the differentiation of BM-MSCs to ECs. CDC42 binds to PAK1, and PAK1 binds to paxillin. In addition, CDC42 in the VEGFA group had a greater ability to bind to PAK1, whereas PAK1 in the VEGFA group had a greater ability to bind to paxillin. Overexpression of miR-206 in VEGFA-induced BM-MSCs demonstrated that MALAT1 competes with the CDC42 3'-UTR for binding to miR-206, which in turn is involved in the differentiation of BM-MSCs to ECs. Compared to the DMED model group, the ICP/MAP ratio was significantly greater in the three BM-MSCs treatment groups. CONCLUSIONS: MALAT1 facilitates BM-MSC differentiation into ECs by regulating the miR-206/CDC42/PAK1/paxillin axis to improve ED. The present findings revealed the vital role of MALAT1 in the repair of BM-MSCs for erectile function and provided new mechanistic insights into the BM-MSC-mediated repair of DMED.

Testis-specific serine kinase 6 (TSSK6) is abnormally expressed in colorectal cancer and promotes oncogenic behaviors.

Cancer testis antigens (CTAs) are a collection of proteins whose expression is normally restricted to the gamete but abnormally activated in a wide variety of tumors. The CTA, Testis-specific serine kinase 6 (TSSK6), is essential for male fertility in mice. The functional relevance of TSSK6 to cancer, if any, has not previously been investigated. Here we find that TSSK6 is frequently anomalously expressed in colorectal cancer and patients with elevated TSSK6 expression have reduced relapse-free survival. Depletion of TSSK6 from colorectal cancer cells attenuates anchorage-independent growth, invasion, and growth in vivo. Conversely, overexpression of TSSK6 enhances anchorage independence and invasion in vitro as well as in vivo tumor growth. Notably, ectopic expression of TSSK6 in semi-transformed human colonic epithelial cells is sufficient to confer anchorage independence and enhance invasion. In somatic cells, TSSK6 co-localizes with and enhances the formation of paxillin and tensin-positive foci at the cell periphery, suggesting a function in focal adhesion formation. Importantly, TSSK6 kinase activity is essential to induce these tumorigenic behaviors. Our findings establish that TSSK6 exhibits oncogenic activity when abnormally expressed in colorectal cancer cells. Thus, TSSK6 is a previously unrecognized intervention target for therapy, which could exhibit an exceptionally broad therapeutic window.

Paxillin regulates androgen receptor expression associated with granulosa cell focal adhesions.

Paxillin is a ubiquitously expressed adaptor protein integral to focal adhesions, cell motility, and apoptosis. Paxillin has also recently been implicated as a mediator of nongenomic androgen receptor (AR) signaling in prostate cancer and other cells. We sought to investigate the relationship between paxillin and AR in granulosa cells (GCs), where androgen actions, apoptosis, and focal adhesions are of known importance, but where the role of paxillin is understudied. We recently showed that paxillin knockout in mouse GCs increases fertility in older mice. Here, we demonstrate that paxillin knockdown in human granulosa-derived KGN cells, as well as knockout in mouse primary GCs, results in reduced AR protein but not reduced mRNA expression. Further, we find that both AR protein and mRNA half-lives are reduced by approximately one-third in the absence of paxillin, but that cells adapt to chronic loss of paxillin by upregulating AR gene expression. Using co-immunofluorescence and proximity ligation assays, we show that paxillin and AR co-localize at the plasma membrane in GCs in a focal adhesion kinase-dependent way, and that disruption of focal adhesions leads to reduced AR protein level. Our findings suggest that paxillin recruits AR to the GC membrane, where it may be sequestered from proteasomal degradation and poised for nongenomic signaling, as reported in other tissues. To investigate the physiological significance of this in disorders of androgen excess, we tested the effect of GC-specific paxillin knockout in a mouse model of polycystic ovary syndrome (PCOS) induced by chronic postnatal dihydrotestosterone (DHT) exposure. While none of the control mice had estrous cycles, 33% of paxillin knockout mice were cycling, indicating that paxillin deletion may offer partial protection from the negative effects of androgen excess by reducing AR expression. Paxillin-knockout GCs from mice with DHT-induced PCOS also produced more estradiol than GCs from littermate controls. Thus, paxillin may be a novel target in the management of androgen-related disorders in women, such as PCOS.

Biological evaluation of sulfonate and sulfate analogues of lithocholic acid: A bioisosterism-guided approach towards the discovery of potential sialyltransferase inhibitors for antimetastatic study.

The naturally occurring bile acid lithocholic acid (LCA) has been a crucial core structure for many non-sugar-containing sialyltranferase (ST) inhibitors documented in literature. With the aim of elucidating the impact of the terminal carboxyl acid substituent of LCA on its ST inhibition, in this present study, we report the (bio)isosteric replacement-based design and synthesis of sulfonate and sulfate analogues of LCA. Among these compounds, the sulfate analogue SPP-002 was found to selectively inhibit N-glycan sialylation by at least an order of magnitude, indicating a substantial improvement in both potency and selectivity when compared to the unmodified parent bile acid. Molecular docking analysis supported the stronger binding of the synthetic analogue in the enzyme active site. Treatment with SPP-002 also hampered the migration, adhesion, and invasion of MDA-MB-231 cells in vitro by suppressing the expression of signaling proteins involved in the cancer metastasis-associated integrin/FAK/paxillin pathway. In totality, these findings offer not only a novel structural scaffold but also valuable insights for the future development of more potent and selective ST inhibitors with potential therapeutic effects against tumor cancer metastasis.

Progerin Inhibits the Proliferation and Migration of Melanoma Cells by Regulating the Expression of Paxillin.

Objective: Progerin, the underlying cause of Hutchinson-Gilford Progeria Syndrome (HGPS), has been extensively studied for its impact on normal cells and premature aging patients. However, there is a lack of research on its specific effects on tumor cells. Melanoma is one of the most common malignant tumors with high morbidity and mortality. This study aimed to elucidate the potential therapeutic role of progerin in melanoma. Materials and Methods: We constructed the melanoma A375 cell line and M14 cell line with stable expression of progerin. The expression of progerin, paxillin, and epithelial-mesenchymal transition (EMT) marker proteins in each cell group was measured using Western blot. The migration, proliferation, and cell cycle of cancer cells were assessed using the transwell assay, wound healing assay, colony formation assay, CCK 8 assay, and flow cytometry. RT-qPCR technology was used to examine the impact of progerin overexpression on microRNA expression. Finally, we transfected paxillin into the progerin overexpression cell group to verify whether progerin regulates the phenotype of tumor cells through paxillin. Results: Our study demonstrated that overexpression of progerin leads to decreased expression of paxillin and inhibits cancer cell migration, proliferation, EMT process and cell cycle progression. Additionally, rescue experiments revealed that the migration, proliferation ability, and EMT marker protein expression in progerin overexpressing cancer cells could be partially restored by transfecting a plasmid containing the paxillin gene. Mechanistic investigations further revealed that progerin achieves this inhibition of paxillin expression by upregulating miR-212. Conclusion: This study reveals that progerin may inhibit the migration and proliferation of melanoma cells through the miR-212/paxillin axis, which provides a new approach for the future treatment of this disease.

Role of steroid hormones in the maintenance of focal adhesions in bovine oviductal epithelial cells.

The oviduct, the organ of the female reproductive system where fertilization and early embryonic development occur, provides an optimal environment for the final maturation of oocytes, storage, and sperm capacitation and transport of gametes and embryos. During the estrous cycle, the oviduct is affected by ovarian sex hormones, resulting in changes aimed at maintaining an appropriate microenvironment. Normal cell migration is tightly regulated, its role being essential for the development and maintenance of organ and tissue functions as well as for regeneration following injury. Due to their involvement in focal contact formations, focal adhesion kinase (PTK2) and paxillin (PXN) are key proteins in the study of cell migration and adhesion. The objective of this work was to compare the expression of PTK2 and PXN in oviductal cells along the estrous cycle and to determine if their expression is regulated by the presence of 17-ß estradiol (E2) and/or progesterone (P4). No transcripts of PTK2 or of PXN were detected in cells corresponding to the luteal phase. Additionally, hormonal stimulation experiments on bovine oviductal cell cultures (BOECs) were carried out, where P4 inhibited the expression of both genes. Migration assays demonstrated that P4 reduced BOECs migration capacity. P4 treatment also reduced cell adhesion, while E2 increased the number of adhered cells. In conclusion, the presence of E2 and P4 regulates the expression of genes involved in the formation of focal contacts and modifies the migration and adhesion of BOECs. Understanding the effect of steroid hormones on BOECs is critical to grasp the impact of steroid control on oviductal function and its contribution to establishing successful pregnancies.

Effect of Paxillin Expression and Phosphorylation on Colorectal Cancer Prognosis and Metastasis.

BACKGROUND/AIM: Colorectal cancer (CRC) is the third most common cancer worldwide, and metastasis is strongly associated with poor prognosis in patients with CRC. We have previously found that the expression and phosphorylation of paxillin (PXN) play an important role in the metastatic potential of breast cancer. This study examined the potential role of PXN in CRC metastasis. MATERIALS AND METHODS: Resected tumor specimens from 92 patients with CRC were subjected to immunohistochemical analysis of PXN levels. Three human CRC cell lines, HCT116, LoVo, and SW480 were used for scratch and transwell invasion assays to examine the effects of PXN over-expression. RNA sequencing was performed to obtain the expression profiles under PXN over-expression. RESULTS: High levels of PXN were significantly correlated with advanced stage, higher carcinoembryonic antigen and carbohydrate antigen 19-9 levels, and poorer overall survival. The migration ability of CRC cells was enhanced by exogenous PXN over-expression, but this enhancement was not observed in cells harboring exogenously mutated PXN at Tyr31 or Tyr88 phosphorylation sites. In PXN-over-expressing cells, TNF-α signaling via NF-[Formula: see text]B was positively enriched. CONCLUSION: PXN expression and phosphorylation at Tyr31 or Tyr88 may influence the migration and invasion of CRC cells. PXN expression and phosphorylation at Tyr31 or Tyr88 are promising targets for evaluating prognosis and treating CRC.

Hic-5 regulates extracellular matrix-associated gene expression and cytokine secretion in cancer associated fibroblasts.

The focal adhesion protein, Hic-5 plays a key role in promoting extracellular matrix deposition and remodeling by cancer associated fibroblasts within the tumor stroma to promote breast tumor cell invasion. However, whether stromal matrix gene expression is regulated by Hic-5 is still unknown. Utilizing a constitutive Hic-5 knockout, Mouse Mammary Tumor Virus-Polyoma Middle T-Antigen spontaneous breast tumor mouse model, bulk RNAseq analysis was performed on cancer associated fibroblasts isolated from Hic-5 knockout mammary tumors. Functional network analysis highlighted a key role for Hic-5 in extracellular matrix organization, with both structural matrix genes, as well as matrix remodeling genes being differentially expressed in relation to Hic-5 expression. The subcellular distribution of the MRTF-A transcription factor and expression of a subset of MRTF-A responsive genes was also impacted by Hic-5 expression. Additionally, cytokine array analysis of conditioned media from the Hic-5 and Hic-5 knockout cancer associated fibroblasts revealed that Hic-5 is important for the secretion of several key factors that are associated with matrix remodeling, angiogenesis and immune evasion. Together, these data provide further evidence of a central role for Hic-5 expression in cancer associated fibroblasts in regulating the composition and organization of the tumor stroma microenvironment to promote breast tumor progression.

Fyn-Mediated Paxillin Tyrosine 31 Phosphorylation Regulates Migration and Invasion of Breast Cancer Cells.

Metastasis is the leading cause of death in breast cancer patients due to the lack of effective therapies. Elevated levels of paxillin expression have been observed in various cancer types, with tyrosine phosphorylation shown to play a critical role in driving cancer cell migration. However, the specific impact of the distinct tyrosine phosphorylation events of paxillin in the progression of breast cancer remains to be fully elucidated. Here, we found that paxillin overexpression in breast cancer tissue is associated with a patient's poor prognosis. Paxillin knockdown inhibited the migration and invasion of breast cancer cells. Furthermore, the phosphorylation of paxillin tyrosine residue 31 (Tyr31) was significantly increased upon the TGF-ß1-induced migration and invasion of breast cancer cells. Inhibiting Fyn activity or silencing Fyn decreases paxillin Tyr31 phosphorylation. The wild-type and constitutively active Fyn directly phosphorylate paxillin Tyr31 in an in vitro system, indicating that Fyn directly phosphorylates paxillin Tyr31. Additionally, the non-phosphorylatable mutant of paxillin at Tyr31 reduces actin stress fiber formation, migration, and invasion of breast cancer cells. Taken together, our results provide direct evidence that Fyn-mediated paxillin Tyr31 phosphorylation is required for breast cancer migration and invasion, suggesting that targeting paxillin Tyr31 phosphorylation could be a potential therapeutic strategy for mitigating breast cancer metastasis.

An ULK1/2-PXN mechanotransduction pathway suppresses breast cancer cell migration.

The remodeling and stiffening of the extracellular matrix (ECM) is a well-recognized modulator of breast cancer progression. How changes in the mechanical properties of the ECM are converted into biochemical signals that direct tumor cell migration and metastasis remain poorly characterized. Here, we describe a new role for the autophagy-inducing serine/threonine kinases ULK1 and ULK2 in mechanotransduction. We show that ULK1/2 activity inhibits the assembly of actin stress fibers and focal adhesions (FAs) and as a consequence impedes cell contraction and migration, independent of its role in autophagy. Mechanistically, we identify PXN/paxillin, a key component of the mechanotransducing machinery, as a direct binding partner and substrate of ULK1/2. ULK-mediated phosphorylation of PXN at S32 and S119 weakens homotypic interactions and liquid-liquid phase separation of PXN, impairing FA assembly, which in turn alters the mechanical properties of breast cancer cells and their response to mechanical stimuli. ULK1/2 and the well-characterized PXN regulator, FAK/Src, have opposing functions on mechanotransduction and compete for phosphorylation of adjacent serine and tyrosine residues. Taken together, our study reveals ULK1/2 as important regulator of PXN-dependent mechanotransduction.

Evaluation of SgK269 expression in colon cancer patients and the effects of hAMSCs secretome on tumor invasion through SgK269/c-Src/p-P130Cas/p-Paxillin/p-ERK1/2 signaling pathway in HT-29 colon cancer cells.

Colon cancer is the fifth leading cause of cancer-related deaths worldwide. Stem cells have unique characteristics and are considered as a novel therapeutic platform for cancer. Sugen Kinase 269 (SgK269) is considered as an oncogenic scaffolding pseudo kinase which governs the rearranging of the cytoskeleton, cellular motility, and invasion. The aim of this study is to evaluate the expression of SgK269 in colon cancer patients and explore the therapeutic effects of human amniotic mesenchymal stromal cells (hAMSCs) on invasion and proliferation of colon cancer cells (HT-29) through analyzing SgK269/c-Src/p-P130Cas/p-Paxillin/p-ERK1/2 signaling pathway. In this regard, we collected 30 samples from colon cancer patients and evaluated SgK269 expression using quantitative real-time PCR (qRT-PCR). Next, we employed a co-culture system using Transwell 6-well plates and after 72 h, tumor growth promotion and invasion were analyzed in hAMSCs-treated HT-29 cells through SgK269/c-Src/p-P130Cas/p-Paxillin/p-ERK1/2/Rac signaling pathway using qRT-PCR, western blot method, MTT assay, wound healing assay, and DAPI staining. Our results showed upregulation of SgK269 in colon cancer patients. Treatment of HT-29 colon cancer cells with hAMSCs secretome can inhibit SgK269/c-Src/p-P130Cas/p-Paxillin/p-ERK1/2/Rac signaling pathway and the resulting suppression of cell invasion and proliferation. Our results suggest that SgK269 is an important target in colon cancer therapy and MSCs secretome may be an effective therapeutic approach to inhibit colon cancer cell invasion and proliferation through SgK269/c-Src/p-P130Cas/p-Paxillin/p-ERK1/2/Rac signaling pathway.

Equine ß-defensin 1 regulates cytokine expression and phagocytosis in S. aureus-infected mouse monocyte macrophages via the Paxillin-FAK-PI3K pathway.

ß-defensin-1 (BD-1) is a rich source of disulfide bonds and antibacterial peptides that exhibit direct bactericidal function. The expression of BD-1 is primarily induced by external stimulation and is known to correlate with TLR-mediated inflammation, suggesting its association with innate immune responses. Equine ß-defensin-1 (eBD-1) belongs to the BD-1 family. Our previous study demonstrated that eBD-1 enhances cytokine expression and promotes macrophage phagocytosis of S. aureus, although the underlying mechanism remains unknown. In this study, we utilized a PI-3K inhibitor (PKI-402) to treat eBD-1 -treated S. aureus-infected macrophages in vitro. Our results revealed that PKI-402 decreased the expression of eBD-1-promoted TNF-α, IL-6, CXCL10, CD40, RANTES, and p65 mRNA. To further investigate the relationship between eBD-1 and phagocytosis, we examined the expression of paxillin and FcγRIII (CD16 receptor) using western blot and immunofluorescence techniques. Our findings demonstrated that eBD-1 enhanced CD16 and paxillin expression in S. aureus -infected macrophages. Considering the correlation between paxillin expression and focal adhesion kinase (FAK), we transfected FAK siRNA into macrophages and evaluated paxillin expression using western blot analysis. Additionally, we quantified the number of S. aureus phagocytosed by macrophages. The results indicated a reduction in both paxillin expression and the number of S. aureus phagocytosed by macrophages upon FAK siRNA treatment. Our study showed the eBD-1 promotes cytokine mRNA expression in S. aureus-infected macrophages regulated by PI-3K-NF-κB pathway, and it increases macrophage phagocytosis of S. aureus associated with the FAK-paxillin signaling pathway.

Evaluation of Paxillin Expression in Epithelial Dysplasia, Oral Squamous Cell Carcinoma, Lichen Planus with and without Dysplasia, and Hyperkeratosis: A Retrospective Cross-Sectional Study.

BACKGROUND: Paxillin is a cytoskeletal protein involved in the pathogenesis of several types of cancers. However, the roles of paxillin in epithelial dysplasia (ED), oral squamous cell carcinoma (OSCC), oral lichen planus with dysplasia (OLPD), hyperkeratosis (HK), and oral lichen planus (OLP) have remained unnoticed in the literature. This study aimed to evaluate its attainable functions in the pathogenesis and malignant transformation of potentially malignant oral epithelium and benign lesions. METHODS: In this retrospective cross-sectional study, paxillin expression was investigated in 99 tissue samples, including 18 cases of OSCC, 21 ED, 23 OLP, 21 OLPD, and 16 cases of HK. The tissue sections also underwent immunohistochemical paxillin staining using 3,3-diaminobenzidine (DAB) chromogen. The intensity, location, and percentage of staining were examined across all groups. Data were analyzed using the Shapiro-Wilk test, ANOVA, Pearson chi-square, Kruskal-Wallis, and Dunn's post hoc test. RESULTS: The cytoplasmic percentage and intensity staining of Paxillin expression were evident in the central/suprabasal and basal/peripheral layers of all the obtained samples. The final staining score was significantly higher in OSCC and dysplasia compared to HK and OLP (p = 0.004). It was found that paxillin expression is associated with the grade of dysplastic samples (p < 0.001). CONCLUSION: The present study provides evidence that paxillin may be involved in the pathogenesis of OSCC and the development and progression of dysplastic tissue, since the paxillin expression was higher than that of HK and OLP.

Epithelial multicellular clustering enabled by polarized macrophages on soft matrices.

Formation of epithelial structures of variegated geometries and sizes is essential for organogenesis, tumor growth, and wound repair. Although epithelial cells are predisposed with potential for multicellular clustering, it remains unclear whether immune cells and mechanical cues from their microenvironment influence this process. To explore this possibility, we cocultured human mammary epithelial cells with prepolarized macrophages on soft or stiff hydrogels. In the presence of M1 (proinflammatory) macrophages on soft matrices, epithelial cells migrated faster and subsequently formed larger multicellular clusters compared to cocultures with M0 (unpolarized) or M2 (anti-inflammatory) macrophages. By contrast, stiff matrices disabled active clustering of epithelial cells due to their enhanced migration and cell-ECM adhesion, regardless of macrophage polarization. We found that the copresence of soft matrices and M1 macrophages reduced focal adhesions, but enhanced fibronectin deposition and nonmuscle myosin-IIA expression, which altogether optimize conditions for epithelial clustering. Upon ROCK inhibition, epithelial clustering was abrogated, indicating a requirement for optimized cellular forces. In these cocultures, TNF-α secretion was the highest with M1 macrophages and TGF-ß secretion was exclusively detectable in case of M2 macrophages on soft gels, which indicated potential role of macrophage secreted factors in the observed epithelial clustering. Indeed, exogenous addition of TGF-ß promoted epithelial clustering with M1 coculture on soft gels. According to our findings, optimization of both mechanical and immune factors can tune epithelial clustering responses, which could have implications in tumor growth, fibrosis, and would healing.

Cdk4 Regulates Glioblastoma Cell Invasion and Stemness and Is Target of a Notch Inhibitor Plus Resveratrol Combined Treatment.

Glioblastoma multiforme (GBM) is one of the most aggressive types of cancer characterized by poor patient outcomes. To date, it is believed that the major cause of its recurrence and chemoresistance is represented by the enrichment of GBM stem cells (GSCs) sustained by the abnormal activation of a number of signaling pathways. In this study, we found that in GBM cells, treatment with low toxicity doses of the γ-secretase inhibitor RO4929097 (GSI), blocking the Notch pathway activity, in combination with resveratrol (RSV) was able to reverse the basal mesenchymal phenotype to an epithelial-like phenotype, affecting invasion and stemness interplay. The mechanism was dependent on cyclin D1 and cyclin-dependent kinase (CDK4), leading to a reduction of paxillin (Pxn) phosphorylation. Consequently, we discovered the reduced interaction of Pxn with vinculin (Vcl), which, during cell migration, transmits the intracellular forces to the extracellular matrix. The exogenous expression of a constitutively active Cdk4 mutant prevented the RSV + GSI inhibitory effects in GBM cell motility/invasion and augmented the expression of stemness-specific markers, as well as the neurosphere sizes/forming abilities in untreated cells. In conclusion, we propose that Cdk4 is an important regulator of GBM stem-like phenotypes and invasive capacity, highlighting how the combined treatment of Notch inhibitors and RSV could be prospectively implemented in the novel therapeutic strategies to target Cdk4 for these aggressive brain tumors.

Arctigenin hinders the invasion and metastasis of cervical cancer cells via the FAK/paxillin pathway.

Context: Cervical cancer is the most common gynecological pernicious tumor with high morbidity and mortality worldwide, especially in developing countries. Arctigenin (ARG), a nature-derived component, has exhibited anti-tumor activity in various tumors. Objective: To explore the effect of ARG on cervical cancer. Materials and methods: The effect and mechanism of ARG on cervical cancer cells were explored by cell counting kit-8 (CCK-8), flow cytometry, transwell and Western blot assays. Additionally, in vivo experiment was conducted in xenografted mice by immunohistochemistry (IHC), terminal deoxynucleotidyl transferase deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL) and Western blot assays. Results: ARG treatment induced both concentration-dependent and time-dependent reductions in the cell viability of SiHa and HeLa cells with a IC50 value of 9.34 µM and 14.45 µM, respectively. ARG increased the apoptosis rate and the protein levels of cleaved-caspase 3 and E-cadherin, but decreased the invaded cell numbers and the protein levels of Vimentin and N-cadherin in vitro. Mechanically, ARG inhibited the expression of focal adhesion kinase (FAK)/paxillin pathway, which was confirmed by the overexpression of FAK in SiHa cells. The inhibitory role of overexpression of FAK in proliferation and invasion, as well as its promoted role in apoptosis were reversed with ARG treatment. Meanwhile, ARG suppressed growth and metastasis, and enhanced apoptosis in vivo. Consistently, ARG administration reduced the relative protein level of p-FAK/FAK and p-paxillin/paxillin in tumor tissues of xenografted mice. Conclusion: ARG inhibited proliferation, invasion and metastasis, but enhanced apoptosis of cervical cancer via the FAK/paxillin axis.

The Role of Paxillin Aberrant Expression in Cancer and Its Potential as a Target for Cancer Therapy.

Paxillin is a multi-domain adaptor protein. As an important member of focal adhesion (FA) and a participant in regulating cell movement, paxillin plays an important role in physiological processes such as nervous system development, embryonic development, and vascular development. However, increasing evidence suggests that paxillin is aberrantly expressed in many cancers. Many scholars have also recognized that the abnormal expression of paxillin is related to the prognosis, metastases, invasion, survival, angiogenesis, and other aspects of malignant tumors, suggesting that paxillin may be a potential cancer therapeutic target. Therefore, the study of how aberrant paxillin expression affects the process of tumorigenesis and metastasis will help to develop more efficacious antitumor drugs. Herein, we review the structure of paxillin and its function and expression in tumors, paying special attention to the multifaceted effects of paxillin on tumors, the mechanism of tumorigenesis and progression, and its potential role in tumor therapy. We also hope to provide a reference for the clinical prognosis and development of new tumor therapeutic targets.