Role of cytoskeleton, cell proliferation and tight junction in wound healing: New therapeutic strategy

By inhibiting Insulin-Like Growth Factor-1-Receptor (IGF-1R) signaling, Klotho (KL) acts like an aging- and tumor-suppressor. We investigated whether KL impacts the aggressiveness of liposarcomas, in which IGF-1R signaling is frequently upregulated. Indeed, we observed that a higher KL expression in liposarcomas is associated with a better outcome for patients. Moreover, KL is downregulated in dedifferentiated liposarcomas (DDLPS) compared to well-differentiated tumors and adipose tissue. Because DDLPS are high-grade tumors associated with poor prognosis, we examined the potential of KL as a tool for overcoming therapy resistance. First, we confirmed the attenuation of IGF-1-induced calcium (Ca2+)-response and Extracellular signal-Regulated Kinase 1/2 (ERK1/2) phosphorylation in KL-overexpressing human DDLPS cells. KL overexpression also reduced cell proliferation, clonogenicity, and increased apoptosis induced by gemcitabine, thapsigargin, and ABT-737, all of which are counteracted by IGF-1R-dependent signaling and activate Ca2+-dependent endoplasmic reticulum (ER) stress. Then, we monitored cell death and cytosolic Ca2+-responses and demonstrated that KL increases the reticular Ca2+-leakage by maintaining TRPC6 at the ER and opening the translocon. Only the latter is necessary for sensitizing DDLPS cells to reticular stressors. This was associated with ERK1/2 inhibition and could be mimicked with IGF-1R or MEK inhibitors. These observations provide a new therapeutic strategy in the management of DDLPS.

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A critical role of AREG for bleomycin-induced skin fibrosis

10.1101/2021.01.20.427509 ◽

2021 ◽

Author(s):

Mary Yinghua Zhang ◽

Shuyi Fang ◽

Hongyu Gao ◽

Xiaoli Zhang ◽

Dongsheng Gu ◽

...

Keyword(s):

Cell Proliferation ◽

Wound Healing ◽

Oral Mucosa ◽

Healing Process ◽

Critical Role ◽

Skin Fibrosis ◽

Wound Healing Process ◽

Organ Function ◽

Signaling Axis

ABSTRACTWe report our discovery of an important player in the development of skin fibrosis, a hallmark of scleroderma. Scleroderma is a fibrotic disease, affecting 70,000 to 150,000 Americans. Fibrosis is a pathological wound healing process that produces an excessive extracellular matrix to interfere with normal organ function. Fibrosis contributes to nearly half of human mortality. Scleroderma has heterogeneous phenotypes, unpredictable outcomes, no validated biomarkers, and no effective treatment. Thus, strategies to slow down scleroderma progression represent an urgent medical need. While a pathological wound healing process like fibrosis leaves scars and weakens organ function, oral mucosa wound healing is a scarless process. After re-analyses of gene expression datasets from oral mucosa wound healing and skin fibrosis, we discovered that several pathways constitutively activated in skin fibrosis are transiently induced during oral mucosa wound healing process, particularly the amphiregulin (Areg) gene. Areg expression is upregulated ~10 folds 24hrs after oral mucosa wound but reduced to the basal level 3 days later. During bleomycin-induced skin fibrosis, a commonly used mouse model for skin fibrosis, Areg is up-regulated throughout the fibrogenesis and is associated with elevated cell proliferation in the dermis. To demonstrate the role of Areg for skin fibrosis, we used mice with Areg knockout, and found that Areg deficiency essentially prevents bleomycin-induced skin fibrosis. We further determined that bleomycin-induced cell proliferation in the dermis was not observed in the Areg null mice. Furthermore, we found that inhibiting MEK, a downstream signaling effector of Areg, by selumetinib also effectively blocked bleomycin-based skin fibrosis model. Based on these results, we concluded that the Areg-EGFR-MEK signaling axis is critical for skin fibrosis development. Blocking this signaling axis may be effective in treating scleroderma.

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Specific NOTCH1 antibody targets DLL4-induced proliferation, migration, and angiogenesis in NOTCH1-mutated CLL cells

Oncogene ◽

10.1038/s41388-019-1053-6 ◽

2019 ◽

Vol 39 (6) ◽

pp. 1185-1197 ◽

Cited By ~ 5

Author(s):

Mónica López-Guerra ◽

Sílvia Xargay-Torrent ◽

Patricia Fuentes ◽

Jocabed Roldán ◽

Blanca González-Farré ◽

...

Keyword(s):

Cell Proliferation ◽

Notch Signaling ◽

Poor Prognosis ◽

Therapeutic Strategy ◽

Target Genes ◽

Notch Signaling Pathway ◽

Lymphocytic Leukemia ◽

Therapeutic Targeting ◽

Notch Ligand

Abstract Targeting Notch signaling has emerged as a promising therapeutic strategy for chronic lymphocytic leukemia (CLL), particularly in NOTCH1-mutated patients. We provide first evidence that the Notch ligand DLL4 is a potent stimulator of Notch signaling in NOTCH1-mutated CLL cells while increases cell proliferation. Importantly, DLL4 is expressed in histiocytes from the lymph node, both in NOTCH1-mutated and -unmutated cases. We also show that the DLL4-induced activation of the Notch signaling pathway can be efficiently blocked with the specific anti-Notch1 antibody OMP-52M51. Accordingly, OMP-52M51 also reverses Notch-induced MYC, CCND1, and NPM1 gene expression as well as cell proliferation in NOTCH1-mutated CLL cells. In addition, DLL4 stimulation triggers the expression of protumor target genes, such as CXCR4, NRARP, and VEGFA, together with an increase in cell migration and angiogenesis. All these events can be antagonized by OMP-52M51. Collectively, our results emphasize the role of DLL4 stimulation in NOTCH1-mutated CLL and confirm the specific therapeutic targeting of Notch1 as a promising approach for this group of poor prognosis CLL patients.

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LncRNA XIST promotes extracellular matrix synthesis, proliferation and migration by targeting miR-29b-3p/COL1A1 in human skin fibroblasts after thermal injury

Biological Research ◽

10.1186/s40659-019-0260-5 ◽

2019 ◽

Vol 52 (1) ◽

Cited By ~ 4

Author(s):

Wei Cao ◽

Youping Feng

Keyword(s):

Extracellular Matrix ◽

Cell Proliferation ◽

Wound Healing ◽

Western Blot ◽

Thermal Injury ◽

Human Skin Fibroblasts ◽

Cell Proliferation And Migration ◽

And Migration ◽

Proliferation And Migration

Abstract Background Long noncoding RNAs (lncRNAs) have been reported to be associated with dermis process during burn wound healing. This study aimed to investigate the role of lncRNA X-inactive specific transcript (XIST) in human skin fibroblasts (HSF) and extracellular matrix (ECM) as well as the regulatory network of XIST/microRNA-29b-3p (miR-29b-3p)/collagen 1 alpha 1 (COL1A1). Methods The wound samples were collected from 25 patients with deep partial thickness burn at day 5 after burn. The thermal injured model was established using HSF cells. The expressions of XIST, miR-29b-3p and COL1A1 were measured by quantitative real-time polymerase chain reaction and western blot. ECM synthesis, cell proliferation and migration were detected by western blot, cell counting kit-8 and trans-well assays, respectively. The interaction between miR-29b-3p and XIST or COL1A1 was explored by bioinformatics analysis and luciferase reporter assay. Results The expressions of XIST and COL1A1 were enhanced but miR-29b-3p expression was decreased after thermal injury. XIST overexpression promoted ECM synthesis, cell proliferation and migration in thermal injured HSF cells. However, XIST knockdown played an opposite effect. miR-29b-3p overexpression inhibited ECM synthesis, cell proliferation and migration, which was reversed by XIST. COL1A1 silence suppressed ECM synthesis, cell proliferation and migration by miR-29b-3p targeting. Moreover, COL1A1 up-regulation weakened the effect of XIST silence on ECM synthesis and HSF cell function. Conclusion XIST promoted ECM synthesis, cell proliferation and migration by sponging miR-29b-3p and targeting COL1A1 in HSF cells after thermal injury, indicating the promoting role of XIST in wound healing.

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BIOL-07. MIR-212 FUNCTIONS AS A TUMOR SUPPRESSOR GENE IN GROUP 3 MEDULLOBLASTOMA VIA TARGETING NUCLEAR FACTOR I/B (NFIB)

Neuro-Oncology ◽

10.1093/neuonc/noab090.014 ◽

2021 ◽

Vol 23 (Supplement_1) ◽

pp. i4-i4

Author(s):

Naveenkumar Perumal ◽

Ranjana Kanchan ◽

David Doss ◽

Pranita Atri ◽

Ramakanth Chirravuri Venkata ◽

...

Keyword(s):

Cell Proliferation ◽

Wound Healing ◽

Tumor Cell ◽

Colony Formation ◽

Tumor Recurrence ◽

Migration And Invasion ◽

Tumor Cell Proliferation ◽

Group 3

Abstract Medulloblastoma (MB), the most frequent malignant pediatric brain tumor is subdivided into four primary subgroups, i.e. wingless-type (WNT), sonic hedgehog (SHH), group 3, and group 4. Haploinsufficiency of chromosome 17p13.3 and c-myc amplification distinguish high-risk group 3 tumors, which are associated with rapid metastasis, recurrence and early mortality. We sought to identify the role of miR-212, which resides on chromosome 17p13.3, in the pathophysiology of group 3 MB. RNA expression analyses revealed dramatically reduced levels of miR-212 in group 3 tumors and cell lines mainly through epigenetic silencing via histone modifications (deacetylation). Restoring in vitro miR-212 expression reduced tumor cell proliferation, colony formation, wound healing, migration and invasion with decreased p-AKT and p-ERK levels in group 3 MB cell lines. Interestingly, a shift in differential c-myc phosphorylation (from serine-62 to threonine-58) was also discovered with miR-212 expression, resulting in reduced total c-myc levels, concurrent with elevated cellular apoptosis. In turn, pro-apoptotic binding partners of c-myc, i.e. Bin-1 and P19ARF, were upregulated in these cells. These findings were recapitulated in stable inducible miR-212 expressing tumor cells. Using a combination of transcriptomic data and a dual luciferase assay, we isolated an important oncogenic target of miR-212, i.e, NFIB, a nuclear transcription factor implicated in metastasis and recurrence. Increased expression of NFIB was confirmed in group 3 tumors, with poor survival shown in high NFIB-expressing patients. As prior, transient NFIB silencing in vitro reduced not only tumor cell proliferation, colony formation, wound healing, migration and invasion, but also medullosphere formation along with decreased expression of stem cell markers (Nanog, Oct4, Sox2, CD133), confirming its role in tumor recurrence possibly via augmenting tumor stemness. Taken together, these results substantiate the tumor suppressive role of miR-212 in group 3 MB and provide a potential new oncogenic target implicated in tumor recurrence, NFIB.

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A critical role of AREG for bleomycin-induced skin fibrosis

Cell & Bioscience ◽

10.1186/s13578-021-00553-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Mary Yinghua Zhang ◽

Shuyi Fang ◽

Hongyu Gao ◽

Xiaoli Zhang ◽

Dongsheng Gu ◽

...

Keyword(s):

Cell Proliferation ◽

Wound Healing ◽

Oral Mucosa ◽

Healing Process ◽

Critical Role ◽

Skin Fibrosis ◽

Wound Healing Process ◽

Organ Function ◽

Signaling Axis

AbstractWe report our discovery of an important player in the development of skin fibrosis, a hallmark of scleroderma. Scleroderma is a fibrotic disease, affecting 70,000 to 150,000 Americans. Fibrosis is a pathological wound healing process that produces an excessive extracellular matrix to interfere with normal organ function. Fibrosis contributes to nearly half of human mortality. Scleroderma has heterogeneous phenotypes, unpredictable outcomes, no validated biomarkers, and no effective treatment. Thus, strategies to slow down scleroderma progression represent an urgent medical need. While a pathological wound healing process like fibrosis leaves scars and weakens organ function, oral mucosa wound healing is a scarless process. After re-analyses of gene expression datasets from oral mucosa wound healing and skin fibrosis, we discovered that several pathways constitutively activated in skin fibrosis are transiently induced during oral mucosa wound healing process, particularly the amphiregulin (Areg) gene. Areg expression is upregulated ~ 10 folds 24hrs after oral mucosa wound but reduced to the basal level 3 days later. During bleomycin-induced skin fibrosis, a commonly used mouse model for skin fibrosis, Areg is up-regulated throughout the fibrogenesis and is associated with elevated cell proliferation in the dermis. To demonstrate the role of Areg for skin fibrosis, we used mice with Areg knockout, and found that Areg deficiency essentially prevents bleomycin-induced skin fibrosis. We further determined that bleomycin-induced cell proliferation in the dermis was not observed in the Areg null mice. Furthermore, we found that inhibiting MEK, a downstream signaling effector of Areg, by selumetinib also effectively blocked bleomycin-based skin fibrosis model. Based on these results, we concluded that the Areg-EGFR-MEK signaling axis is critical for skin fibrosis development. Blocking this signaling axis may be effective in treating scleroderma.

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Oncogenic role of MiR-130a in oral squamous cell carcinoma

Scientific Reports ◽

10.1038/s41598-021-87388-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Karthik Mallela ◽

Swamy Shivananda ◽

Kodaganur S. Gopinath ◽

Arun Kumar

Keyword(s):

Cell Proliferation ◽

Squamous Cell Carcinoma ◽

Oral Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Therapeutic Strategy ◽

Inverse Correlation ◽

Migration And Invasion ◽

Cellular Functions

AbstractAberrant activation of the PI3K/AKT/mTOR pathway is attributed to the pathogenesis of oral squamous cell carcinoma (OSCC). In recent years, increasing evidence suggests the involvement of microRNAs (miRNAs) in oral carcinogenesis by acting as tumor suppressors or oncogenes. TSC1, as a component of the above pathway, regulates several cellular functions such as cell proliferation, apoptosis, migration and invasion. Downregulation of TSC1 is reported in oral as well as several other cancers and is associated with an unfavourable clinical outcome in patients. Here we show that oncogenic miR-130a binds to the 3′UTR of TSC1 and represses its expression. MiR-130a-mediated repression of TSC1 increases cell proliferation, anchorage independent growth and invasion of OSCC cells, which is dependent on the presence of the 3′UTR in TSC1. We observe an inverse correlation between the expression levels of miR-130a and TSC1 in OSCC samples, suggesting that their interaction is physiologically relevant. Delivery of antagomiR-130a to OSCC cells results in a significant decrease in xenograft size. Taken together, the findings of the study indicate that miR-130a-mediated TSC1 downregulation is not only a novel mechanism in OSCC, but also the restoration of TSC1 levels by antagomiR-130a may be a potential therapeutic strategy for the treatment of OSCC.

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Oncogenic Role of miR-130a in Oral Squamous Cell Carcinoma

10.21203/rs.3.rs-130377/v1 ◽

2020 ◽

Author(s):

Karthik Mallela ◽

Shivananda Swamy ◽

K. Gopinath ◽

Arun Kumar

Keyword(s):

Cell Proliferation ◽

Squamous Cell Carcinoma ◽

Oral Squamous Cell Carcinoma ◽

Cell Carcinoma ◽

Squamous Cell ◽

Therapeutic Strategy ◽

Inverse Correlation ◽

Migration And Invasion ◽

Cellular Functions

Abstract Aberrant activation of the PI3K/AKT/mTOR pathway is attributed to the pathogenesis of oral squamous cell carcinoma (OSCC). In recent years, increasing evidence suggests the involvement of microRNAs (miRNAs) in oral carcinogenesis by acting as tumor suppressors or oncogenes. TSC1, as a component of the above pathway, regulates several cellular functions such as cell proliferation, apoptosis, migration and invasion. Downregulation of TSC1 is reported in oral as well as several other cancers and is associated with an unfavourable clinical outcome in patients. Here we show that oncogenic miR-130a binds to the 3’UTR of TSC1 and represses its expression. MiR-130a-mediated repression of TSC1 increases cell proliferation, anchorage independent growth and invasion of OSCC cells, which is dependent on the presence of the 3’UTR in TSC1. We observe an inverse correlation between the expression levels of miR-130a and TSC1 in OSCC samples, suggesting that their interaction is physiologically relevant. Delivery of antagomiR-130a to OSCC cells results in a significant decrease in xenograft size. Taken together, the findings of the study indicate that miR-130a-mediated TSC1 downregulation is not only a novel mechanism in OSCC, but the restoration of TSC1 levels by antagomiR-130a may be a potential therapeutic strategy for the treatment of OSCC.

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Abstract 992: Novel Role of Copper-dependent Transcription Factor Antioxidant-1 in Cell Proliferation and Wound Healing

Circulation ◽

10.1161/circ.116.suppl_16.ii_197 ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Ha Won Kim ◽

Kiyoshi Ozumi ◽

McKinney D Ronald ◽

Shinichi Itoh ◽

Jay Yang ◽

...

Keyword(s):

Cell Proliferation ◽

Wound Healing ◽

Transcription Factor ◽

Cyclin D1 ◽

Wound Closure ◽

Copper Chaperone ◽

Dependent Manner ◽

Chaperone Function

Copper plays a fundamental role in regulating cell proliferation involved in wound healing and angiogenesis. Recently, we found that antioxidant-1 (Atox1), previously appreciated as a copper chaperone at the transGolgi network (TGN), stimulates cyclin D1 transcription and cell proliferation in a copper dependent manner. However, functional role of Atox1 in wound healing in vivo remains unknown. Here we show that Atox1 was highly expressed at the BrdU positive nuclei in the wounding tissue, which was associated with an increase in copper content (2.1-fold), as assessed by ICP-mass spectrometry, as compared to non-wounded skin. In Atox1 knockout (KO) mice, the rate of wound closure was markedly delayed as compared to wild-type (WT) mice (7.1 day vs. 11.2 day for complete wound closure in WT vs. Atox1 KO mice, p < 0.001), and BrdU-positive proliferating cells and cyclin D1 protein expression were significantly decreased. Capillary density in healing wound tissue was also reduced in Atox1 KO mice (27.8% decrease). To segregate a copper-dependent transcription factor and a copper chaperone function of Atox1, we have generated lentivirus expressing nuclear targeted Atox1 (Atox1-NLS), TGN-targeted Atox1 (Atox1-TGN) or Atox1-WT. and examine their effect on cell proliferation and wound healing in vitro and in vivo. In cultured Atox1 KO mouse fibroblasts, copper-induced cell proliferation and cyclin D1 expression were abolished, which was rescued by re-expression of both Atox1-NLS and Atox1-WT, but not Atox1-TGN. Promoter deletion analysis identified Atox1 responsive element (Atox1-RE) at cyclin D1 promoter and EMSA and ChIP assays demonstrated that Atox1 binds to Atox1-RE at the cyclin D1 promoter in a copper dependent manner. The impairment of wound healing in Atox1 KO mice was rescued by re-expression with lenti-Atox1-NLS and Atox1-WT, but not Atox1-TGN, which was associated with the increase in cyclin D1 expression as well as number of BrdU positive proliferating nuclei. In summary, transcription factor function of Atox1, but not copper-chaperone function of Atox1, plays a critical role in copper-induced cell proliferation, which may contribute to wound repair process in response to injury in vivo.

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Impact of KLF4 on Cell Proliferation and Epithelial Differentiation in the Context of Cystic Fibrosis

International Journal of Molecular Sciences ◽

10.3390/ijms21186717 ◽

2020 ◽

Vol 21 (18) ◽

pp. 6717

Author(s):

Luís Sousa ◽

Ines Pankonien ◽

Filipa B. Simões ◽

Marc Chanson ◽

Margarida D. Amaral

Keyword(s):

Cystic Fibrosis ◽

Cell Proliferation ◽

Wound Healing ◽

Epithelial Differentiation ◽

Transepithelial Electrical Resistance ◽

Negative Regulator ◽

Differential Impact ◽

Differentiation Markers ◽

A Cell

Cystic fibrosis (CF) cells display a more cancer-like phenotype vs. non-CF cells. KLF4 overexpression has been described in CF and this transcriptional factor acts as a negative regulator of wt-CFTR. KLF4 is described as exerting its effects in a cell-context-dependent fashion, but it is generally considered a major regulator of proliferation, differentiation, and wound healing, all the processes that are also altered in CF. Therefore, it is relevant to characterize the differential role of KLF4 in these processes in CF vs. non-CF cells. To this end, we used wt- and F508del-CFTR CFBE cells and their respective KLF4 knockout (KO) counterparts to evaluate processes like cell proliferation, polarization, and wound healing, as well as to compare the expression of several epithelial differentiation markers. Our data indicate no major impact of KLF4 KO in proliferation and a differential impact of KLF4 KO in transepithelial electrical resistance (TEER) acquisition and wound healing in wt- vs. F508del-CFTR cells. In parallel, we also observed a differential impact on the levels of some differentiation markers and epithelial-mesencymal transition (EMT)-associated transcription factors. In conclusion, KLF4 impacts TEER acquisition, wound healing, and the expression of differentiation markers in a way that is partially dependent on the CFTR-status of the cell.

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