Connective Tissue Growth Factor: Regulation, Diseases, and Drug Discovery.

In drug discovery, selecting targeted molecules is crucial as the target could directly affect drug efficacy and the treatment outcomes. As a member of the CCN family, CTGF (also known as CCN2) is an essential regulator in the progression of various diseases, including fibrosis, cancer, neurological disorders, and eye diseases. Understanding the regulatory mechanisms of CTGF in different diseases may contribute to the discovery of novel drug candidates. Summarizing the CTGF-targeting and -inhibitory drugs is also beneficial for the analysis of the efficacy, applications, and limitations of these drugs in different disease models. Therefore, we reviewed the CTGF structure, the regulatory mechanisms in various diseases, and drug development in order to provide more references for future drug discovery.

The Role of Collagen VIII in the Aging Mouse Kidney.

The gradual loss of kidney function due to increasing age is accompanied by structural changes such as fibrosis of the tissue. The underlying molecular mechanisms are complex, but not yet fully understood. Non-fibrillar collagen type VIII (COL8) could be a potential factor in the fibrosis processes of the aging kidney. A pathophysiological significance of COL8 has already been demonstrated in the context of diabetic kidney disease, with studies showing that it directly influences both the development and progression of renal fibrosis occurring. The aim of this study was to investigate whether COL8 impacts age-related micro-anatomical and functional changes in a mouse model. The kidneys of wild-type (Col8-wt) and COL8-knockout (Col8-ko) mice of different age and sex were characterized with regard to the expression of molecular fibrosis markers, the development of nephrosclerosis and renal function. The age-dependent regulation of COL8 mRNA expression in the wild-type revealed sex-dependent effects that were not observed with collagen IV (COL4). Histochemical staining and protein analysis of profibrotic cytokines TGF-ß1 (transforming growth factor) and CTGF (connective tissue growth factor) in mouse kidneys showed significant age effects as well as interactions of the factors age, sex and Col8 genotype. There were also significant age and Col8 genotype effects in the renal function data analyzed by urinary cystatin C. In summary, the present study shows, for the first time, that COL8 is regulated in an age- and sex-dependent manner in the mouse kidney and that the expression of COL8 influences the severity of age-induced renal fibrosis and function.

Linagliptin ameliorates tacrolimus-induced renal injury: role of Nrf2/HO-1 and HIF-1α/CTGF/PAI-1.

BACKGROUND: Tacrolimus (TAC) is a frequently used immunosuppressive medication in organ transplantation. However, its nephrotoxic impact limits its long-term usage. This study aims to investigate the effect of linagliptin (Lina) on TAC-induced renal injury and its underlying mechanisms. METHODS AND RESULTS: Thirty-two Sprague Dawley rats were treated with TAC (1.5 mg/kg/day, subcutaneously) and/or Lina (5 mg/kg/day, orally) for 4 weeks. Histological examination was conducted, and serum and urinary biomarkers were measured to assess kidney function and integrity. Furthermore, ELISA, Western blot analysis and immunohistochemical assay were employed to determine signaling molecules of oxidative stress, profibrogenic, hypoxic, and apoptotic proteins. Tacrolimus caused renal dysfunction and histological deterioration evidenced by increased serum creatinine, blood urea nitrogen (BUN), urinary cystatin C, and decreased serum albumin as well as elevated tubular injury and interstitial fibrosis scores. Additionally, TAC significantly increased the expression of collagen type-1, alpha-smooth muscle actin (α-SMA), plasminogen activator inhibitor-1 (PAI-1), and transforming growth factor-beta1 (TGF-ß1) renal content. Moreover, TAC decreased the expression of nuclear factor erythroid-2-related factor2 (Nrf2), heme oxygenase 1 (HO-1), and mitochondrial superoxide dismutase (SOD2). In addition, TAC increased protein expression of hypoxia-inducible factor1-alpha (HIF-1α), connective tissue growth factor (CTGF), inducible nitric oxide synthase (iNOS), 8-hydroxy-2-deoxyguanosine (8-OHdG), as well as nitric oxide (NO), 4-hydroxynonenal, caspase-3 and Bax renal contents. Furthermore, TAC decreased Bcl-2 renal contents. The Lina administration markedly attenuated these alterations. CONCLUSION: Lina ameliorated TAC-induced kidney injury through modulation of oxidative stress, hypoxia, and apoptosis related proteins.

Lipid nephrotoxicity mediated by HIF-1α activation accelerates tubular injury in diabetic nephropathy.

This study is intended to explore the effect of hypoxia-inducible factor-1α (HIF-1α) activation on lipid accumulation in the diabetic kidney. A type 1 diabetic rat model was established by STZ intraperitoneal injection. Cobalt chloride (CoCl2) and YC-1 were used as the HIF-1α activator and antagonist, respectively. CoCl2 treatment significantly increased HIF-1α expression, accelerated lipid deposition, and accelerated tubular injury in diabetic kidneys. In vitro, CoCl2 effectively stabilized HIF-1α and increased its transportation from the cytoplasm to the nucleus, which was accompanied by significantly increased lipid accumulation in HK-2 cells. Furthermore, results obtained in vivo showed that HIF-1α protein expression in the renal tubules of diabetic rats was significantly downregulated by YC-1 treatment. Meanwhile, lipid accumulation in the tubules of the DM + YC-1 group was markedly decreased in comparison to the DM + DMSO group. Accordingly, PAS staining revealed that the pathological injury caused to the tubular epithelial cells was alleviated by YC-1 treatment. Furthermore, the blood glucose level, urine albumin creatinine ratio, and NAG creatinine ratio in the DM + YC-1 group were significantly decreased compared to the DM + DMSO group. Moreover, the protein expression levels of transforming growth factor ß1 (TGF-ß1) and connective tissue growth factor (CTGF) in diabetic kidneys were decreased by YC-1 treatment. Our findings demonstrate that the activation of HIF-1α contributed to interstitial injury in a rat model of diabetic nephropathy and that the underlying mechanism involved the induction of lipid accumulation.

The yes-associated protein-1 (YAP1) inhibitor celastrol suppresses the ability of transforming growth factor ß to activate human gingival fibroblasts.

OBJECTIVE: To determine whether celastrol, an inhibitor of the mechanosensitive transcriptional cofactor yes-associated protein-1 (YAP1), impairs the ability of TGFß1 to stimulate fibrogenic activity in human gingival fibroblast cell line. DESIGN: Human gingival fibroblasts were pre-treated with celastrol or DMSO followed by stimulation with or without TGFß1 (4 ng/ml). We then utilized bulk RNA sequencing (RNAseq), real-time polymerase chain reaction (RT-PCR), Western blot, immunofluorescence, cell proliferation assays to determine if celastrol impaired TGFß1-induced responses in a human gingival fibroblast cell line. RESULTS: Celastrol impaired the ability of TGFß1 to induce expression of the profibrotic marker and mediator CCN2. Bulk RNAseq analysis of gingival fibroblasts treated with TGFß1, in the presence or absence of celastrol, revealed that celastrol impaired the ability of TGFß1 to induce mRNA expression of genes within extracellular matrix, wound healing, focal adhesion and cytokine/Wnt signaling clusters. RT-PCR analysis of extracted RNAs confirmed that celastrol antagonized the ability of TGFß1 to induce expression of genes anticipated to contribute to fibrotic responses. Celastrol also reduced gingival fibroblast proliferation, and YAP1 nuclear localization in response to TGFß1. CONCLUSION: YAP1 inhibitors such as celastrol could be used to impair pro-fibrotic responses to TGFß1 in human gingival fibroblasts.

The association of plasma connective tissue growth factor levels with left ventricular diastolic dysfunction in patients with overt hyperthyroidism.

Background: Left ventricular (LV) diastolic dysfunction is an independent predictor of future cardiovascular events. Early detection of patients with LV diastolic dysfunction can improve clinical outcomes through active management. However, the assessment of diastolic function is very complicated, and there are currently lack of effective biomarkers to assess the risk of LV diastolic dysfunction. Connective tissue growth factor (CTGF) plays a significant role in cardiac remodeling and dysfunction. We aimed to investigate the associations between plasma CTGF level and the risk of LV diastolic dysfunction in this study and judge its effectiveness in diagnosing LV diastolic dysfunction. Methods: A total of 169 patients with overt hyperthyroidism were included. LV diastolic function was evaluated and the subjects were divided into normal LV diastolic function group and LV diastolic dysfunction group. Routine clinical medical data, biochemical data, thyroid related parameters and echocardiographic parameters were recorded for analysis. Results: Compared with normal LV diastolic function group, the LV diastolic dysfunction group had higher age and BMI, as well as lower heart rate, lower serum albumin, lower eGFR, higher serum TgAb and BNP level, and the incidences of hypertension were also higher (all P <0.05). Circulating plasma CTGF levels in the LV diastolic dysfunction group were significantly higher (normal LV diastolic function group: 7.026 [5.567-8.895], LV diastolic dysfunction group: 8.290 [7.054-9.225] ng/ml, median [(Interquartile range)], P = 0.004); Compared with the lowest quartile group, the crude odds ratios (OR) of LV diastolic dysfunction in the second, third, and fourth quartile group were 3.207, 5.032 and 4.554, respectively (all P<0.05). After adjustment for the potentially confounding variables, the adjusted OR values of the third and fourth quartile group had no obvious change. The results of ROC showed that the plasma CTGF had the largest area under the ROC curve, and the value was 0.659 (P = 0.005). Conclusion: The level of circulating plasma CTGF in the LV diastolic dysfunction group was significantly increased. Plasma CTGF level is an independent risk factor for LV diastolic dysfunction. Compared with serum BNP level, the plasma CTGF level may have auxiliary diagnostic value for LV diastolic dysfunction in hyperthyroid patients.

Connective tissue growth factor enhances TGF-ß1-induced osteogenic differentiation via activation of p38 MAPK in mesenchymal stem cells.

OBJECTIVES: Cellular differentiation is based on the effects of various growth factors. Transforming growth factor (TGF)-ß1 plays a pivotal role in inducing osteogenic differentiation of mesenchymal stem cells (MSCs). In this study, we investigated the influence of connective tissue growth factor (CTGF), known to function synergistically with TGF-ß1, on osteogenic differentiation in MSCs. METHODS: UE7T-13 cells were treated with TGF-ß1 and/or CTGF. Subsequently, protein levels of intracellular signaling pathway molecules were determined through western blot analysis. The mRNA expression levels of osteogenic differentiation markers were investigated using reverse transcription-quantitative polymerase chain reaction. Bone matrix mineralization was evaluated through alizarin red staining. RESULTS: Co-treatment with TGF-ß1 and CTGF resulted in the suppression of TGF-ß1-induced phosphorylation of extracellular signal-regulated kinase 1/2, an intracellular signaling pathway molecule in MSCs, while significantly enhancing the phosphorylation of p38 mitogen-activated protein kinase (MAPK). In MSCs, co-treatment with CTGF and TGF-ß1 led to increased expression levels of alkaline phosphatase and type I collagen, markers of osteogenic differentiation induced by TGF-ß1. Osteopontin expression was observed only after TGF-ß1 and CTGF co-treatment. Notably, bone sialoprotein and osteocalcin were significantly upregulated by treatment with CTGF alone. Furthermore, CTGF enhanced the TGF-ß1-induced mineralization in MSCs, with complete suppression observed after treatment with a p38 MAPK inhibitor. CONCLUSIONS: CTGF enhances TGF-ß1-induced osteogenic differentiation and subsequent mineralization in MSCs by predominantly activating the p38 MAPK-dependent pathway.

Serum level of full-length connective tissue growth factor reflects liver fibrosis stage in patients with Fontan-associated liver disease.

BACKGROUND: Chronic liver disease leads to liver fibrosis, and an accurate diagnosis of the fibrosis stage is crucial for medical management. Connective tissue growth factor (CTGF) is produced by endothelial cells and platelets and plays a central role in inducing fibrosis in various organs. In the present study, we tested the validity of measuring the serum levels of two types of CTGF to estimate the biopsy-confirmed liver fibrosis stage. METHODS: We used two detection antibodies targeting the N- and C-terminal of CTGF to measure the serum levels of two forms of CTGF consisting of its full length and its N-terminal fragment. We analyzed the level of CTGF (via enzyme-linked immunosorbent assay) and the liver fibrosis stage in 38 patients with Fontan-associated liver disease (FALD) (26 cases of which were diagnosed pathologically). Correlations were determined by multivariate analysis and the area under the receiver operating characteristic curve. The 65 patients with nonalcoholic fatty liver disease (NAFLD) were included as a disease control group for examination. RESULTS: Full-length CTGF was significantly inversely correlated with liver fibrosis in patients with FALD. Although the platelet count was also associated with the liver fibrosis stage, full-length CTGF was more closely correlated with the fibrosis stage. Furthermore, the level of full-length CTGF was inversely associated with high central venous pressure. Conversely, the serum level of CTGF was not correlated with the fibrosis stage in NAFLD. CONCLUSION: The serum level of full-length CTGF may be useful for estimating the liver fibrosis stage in patients with FALD.

Characterization of gastric cancer-stimulated signaling pathways and function of CTGF in cancer-associated fibroblasts.

BACKGROUND: Cancer-associated fibroblasts (CAFs) are key components of the tumor microenvironment (TME) that play an important role in cancer progression. Although the mechanism by which CAFs promote tumorigenesis has been well investigated, the underlying mechanism of CAFs activation by neighboring cancer cells remains elusive. In this study, we aim to investigate the signaling pathways involved in CAFs activation by gastric cancer cells (GC) and to provide insights into the therapeutic targeting of CAFs for overcoming GC. METHODS: Alteration of receptor tyrosine kinase (RTK) activity in CAFs was analyzed using phospho-RTK array. The expression of CAFs effector genes was determined by RT-qPCR or ELISA. The migration and invasion of GC cells co-cultured with CAFs were examined by transwell migration/invasion assay. RESULTS: We found that conditioned media (CM) from GC cells could activate multiple receptor tyrosine kinase signaling pathways, including ERK, AKT, and STAT3. Phospho-RTK array analysis showed that CM from GC cells activated PDGFR tyrosine phosphorylation, but only AKT activation was PDGFR-dependent. Furthermore, we found that connective tissue growth factor (CTGF), a member of the CCN family, was the most pronouncedly induced CAFs effector gene by GC cells. Knockdown of CTGF impaired the ability of CAFs to promote GC cell migration and invasion. Although the PDGFR-AKT pathway was pronouncedly activated in CAFs stimulated by GC cells, its pharmacological inhibition affected neither CTGF induction nor CAFs-induced GC cell migration. Unexpectedly, the knockdown of SRC and SRC-family kinase inhibitors, dasatinib and saracatinib, significantly impaired CTGF induction in activated CAFs and the migration of GC cells co-cultured with CAFs. SRC inhibitors restored the reduced expression of epithelial markers, E-cadherin and Zonula Occludens-1 (ZO-1), in GC cells co-cultured with CAFs, as well as CAFs-induced aggregate formation in a 3D tumor spheroid model. CONCLUSIONS: This study provides a characterization of the signaling pathways and effector genes involved in CAFs activation, and strategies that could effectively inhibit it in the context of GC. Video Abstract.

Aerobic exercise protects MI heart through miR-133a-3p downregulation of connective tissue growth factor.

OBJECTIVE: To investigate the effect of aerobic exercise intervention to inhibit cardiomyocyte apoptosis and thus improve cardiac function in myocardial infarction (MI) mice by regulating CTGF expression through miR-133a-3p. METHODS: Male C57/BL6 mice, 7-8 weeks old, were randomly divided into sham-operated group (S group), sham-operated +aerobic exercise group (SE group), myocardial infarction group (MI group) and MI + aerobic exercise group (ME group). The mice were anesthetized the day after training and cardiac function was assessed by cardiac echocardiography. Myocardial collagen volume fraction (CVF%) was analyzed by Masson staining. Myocardial CTGF, Bax and Bcl-2 were detected by Western blotting, and myocardial miR-133a-3p was measured by RT-qPCR. RESULTS: Compared with the S group, miR-133a-3p, Bcl-2 and EF were significantly decreased and CTGF, Bax, Bax/ Bcl-2, Caspase 3, Cleaved Caspase-3, LVIDd, LVIDs and CVF were significantly increased in the MI group. Compared with the MI group, miR-133a-3p, Bcl-2 and EF were significantly increased, cardiac function was significantly improved, and CTGF, Bax, Bax/ Bcl-2, Caspase 3, Cleaved Caspase-3, LVIDd, LVIDs and CVF were significantly decreased in ME group. The miR-133a-3p was significantly lower and CTGF was significantly higher in the H2O2 intervention group compared with the control group of H9C2 rat cardiomyocytes. miR-133a-3p was significantly higher and CTGF was significantly lower in the AICAR intervention group compared to the H2O2 intervention group. Compared with the control group of H9C2 rat cardiomyocytes, CTGF, Bax and Bax/Bcl-2 were significantly increased and Bcl-2 was significantly decreased in the miR-133a-3p inhibitor intervention group; CTGF, Bax and Bax/Bcl-2 were significantly decreased and Bcl-2 was significantly upregulated in the miR-133a-3p mimics intervention group. CONCLUSION: Aerobic exercise down-regulated CTGF expression in MI mouse myocardium through miR-133a-3p, thereby inhibiting cardiomyocyte apoptosis and improving cardiac function.

CTGF regulates mineralization in human mature chondrocyte by controlling Pit-1 and modulating ANK via the BMP/Smad signalling.

OBJECTIVE: Connective tissue growth factor (CTGF) exhibits potent proliferative, differentiated, and mineralizing effects, and is believed to be contribute to cartilage mineralization in Osteoarthritis (OA). However, the underlying mechanism of chondrocyte mineralization induced by CTGF remains obscure. As a key regulator of mineral responses, type III phosphate transporter 1 (Pit-1) has been associated with the pathogenesis of articular mineralization. Therefore, the primary objective of this study was to investigate whether CTGF influences the development of mature chondrocyte mineralization and the underlying mechanisms governing such mineralization. METHODS: The effect of Connective tissue growth factor (CTGF) on human C-28/I2 chondrocytes were investigated. The chondrocytes were treated with CTGF or related inhibitors, and transfected with Overexpression and siRNA transfection of Type III Phosphate Transporter 1(Pit-1). Subsequently, the cells were subjected to Alizarin red S staining, PiPer Phosphate Assay Kit, Alkaline Phosphatase Diethanolamine Activity Kit, ELISA, RT-PCR or Western blot analysis. RESULTS: Stimulation with Connective tissue growth factor (CTGF) significantly upregulated the expression of the Type III Phosphate Transporter 1(Pit-1) and mineralization levels in chondrocytes through activation of α5ß1 integrin and BMP/Samd1/5/8 signaling pathways. Furthermore, treatment with overexpressed Pit-1 markedly increased the expression of Multipass Transmembrane Ankylosis (ANK) transporter in the cells. The inhibitory effect of CTGF receptor blockade using α5ß1 Integrin blocking antibody was demonstrated by significantly suppressed the expression of Pit-1 and ANK transporter, as well as chondrocyte mineralization. CONCLUSIONS: Our data indicate that Connective tissue growth factor (CTGF) plays a critical role inchondrocyte mineralization, which is dependent on the expression of the Type III Phosphate Transporter 1(Pit-1) and Multipass Transmembrane Ankylosis (ANK) transporter. Consequently, inhibition of CTGF activity may represent a novel therapeutic approach for the management of Osteoarthritis (OA).

Therapeutic strategies to target connective tissue growth factor in fibrotic lung diseases.

The treatment of interstitial lung diseases, including idiopathic pulmonary fibrosis (IPF), remains challenging as current available antifibrotic agents are not effective in halting disease progression. Connective tissue growth factor (CTGF), also known as cellular communication factor 2 (CCN2), is a member of the CCN family of proteins that regulates cell signaling through cell surface receptors such as integrins, the activity of cytokines/growth factors, and the turnover of extracellular matrix (ECM) proteins. Accumulating evidence indicates that CTGF plays a crucial role in promoting lung fibrosis through multiple processes, including inducing transdifferentiation of fibroblasts to myofibroblasts, epithelial-mesenchymal transition (EMT), and cooperating with other fibrotic mediators such as TGF-ß. Increased expression of CTGF has been observed in fibrotic lungs and inhibiting CTGF signaling has been shown to suppress lung fibrosis in several animal models. Thus, the CTGF signaling pathway is emerging as a potential therapeutic target in IPF and other pulmonary fibrotic conditions. This review provides a comprehensive overview of the current evidence on the pathogenic role of CTGF in pulmonary fibrosis and discusses the current therapeutic agents targeting CTGF using a systematic review approach.

Unraveling Connective Tissue Growth Factor as a Therapeutic Target and Assessing Kahweol as a Potential Drug Candidate in Triple-Negative Breast Cancer Treatment.

Triple-negative breast cancer (TNBC) is characterized by aggressive behavior and limited treatment options, necessitating the identification of novel therapeutic targets. In this study, we investigated the clinical significance of connective tissue growth factor (CTGF) as a prognostic marker and explored the potential therapeutic effects of kahweol, a coffee diterpene molecule, in TNBC treatment. Initially, through a survival analysis on breast cancer patients from The Cancer Genome Atlas (TCGA) database, we found that CTGF exhibited significant prognostic effects exclusively in TNBC patients. To gain mechanistic insights, we performed the functional annotation and gene set enrichment analyses, revealing the involvement of CTGF in migratory pathways relevant to TNBC treatment. Subsequently, in vitro experiments using MDA-MB 231 cells, a representative TNBC cell line, demonstrated that recombinant CTGF (rCTGF) administration enhanced cell motility, whereas CTGF knockdown using CTGF siRNA resulted in reduced motility. Notably, rCTGF restored kahweol-reduced cell motility, providing compelling evidence for the role of CTGF in mediating kahweol's effects. At the molecular level, kahweol downregulated the protein expression of CTGF as well as critical signaling molecules, such as p-ERK, p-P38, p-PI3K/AKT, and p-FAK, associated with cell motility. In summary, our findings propose CTGF as a potential prognostic marker for guiding TNBC treatment and suggest kahweol as a promising antitumor compound capable of regulating CTGF expression to suppress cell motility in TNBC. These insights hold promise for the development of targeted therapies and improved clinical outcomes for TNBC patients.

Blocking CCN2 Reduces Established Palmar Neuromuscular Fibrosis and Improves Function Following Repetitive Overuse Injury.

The matricellular protein cell communication factor 2/connective tissue growth factor (CCN2/CTGF) is critical to development of neuromuscular fibrosis. Here, we tested whether anti-CCN2 antibody treatment will reduce established forepaw fibro-degenerative changes and improve function in a rat model of overuse injury. Adult female rats performed a high repetition high force (HRHF) task for 18 weeks. Tissues were collected from one subset after 18 wks (HRHF-Untreated). Two subsets were provided 6 wks of rest with concurrent treatment with anti-CCN2 (HRHF-Rest/anti-CCN2) or IgG (HRHF-Rest/IgG). Results were compared to IgG-treated Controls. Forepaw muscle fibrosis, neural fibrosis and entheseal damage were increased in HRHF-Untreated rats, compared to Controls, and changes were ameliorated in HRHF-Rest/anti-CCN2 rats. Anti-CCN2 treatment also reduced phosphorylated-ß-catenin (pro-fibrotic protein) in muscles and distal bone/entheses complex, and increased CCN3 (anti-fibrotic) in the same tissues, compared to HRHF-Untreated rats. Grip strength declines and mechanical sensitivity observed in HRHF-Untreated improved with rest; grip strength improved further in HRHF-Rest/anti-CCN2. Grip strength declines correlated with muscle fibrosis, entheseal damage, extraneural fibrosis, and decreased nerve conduction velocity, while enhanced mechanical sensitivity (a pain-related behavior) correlated with extraneural fibrosis. These studies demonstrate that blocking CCN2 signaling reduces established forepaw neuromuscular fibrosis and entheseal damage, which improves forepaw function, following overuse injury.

CTGF promotes the repair and regeneration of alveoli after acute lung injury by promoting the proliferation of subpopulation of AEC2s.

BACKGROUND: Functional alveolar regeneration is essential for the restoration of normal lung homeostasis after acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Lung is a relatively quiescent organ and a variety of stem cells are recruited to participate in lung repair and regeneration after lung tissue injury. However, there is still no effective method for promoting the proliferation of endogenous lung stem cells to promote repair and regeneration. METHODS: Using protein mass spectrometry analysis, we analyzed the microenvironment after acute lung injury. RNA sequencing and image cytometry were used in the alveolar epithelial type 2 cells (AEC2s) subgroup identification. Then we used Sftpc+AEC2 lineage tracking mice and purified AEC2s to further elucidate the molecular mechanism by which CTGF regulates AEC2s proliferation both in vitro and in vivo. Bronchoalveolar lavage fluid (BALF) from thirty ARDS patients who underwent bronchoalveolar lavage was collected for the analysis of the correlation between the expressing of Krt5 in BALF and patients' prognosis. RESULTS: Here, we elucidate that AEC2s are the main facultative stem cells of the distal lung after ALI and ARDS. The increase of connective tissue growth factor (CTGF) in the microenvironment after ALI promoted the proliferation of AEC2s subpopulations. Proliferated AEC2s rapidly expanded and differentiated into alveolar epithelial type 1 cells (AEC1s) in the regeneration after ALI. CTGF initiates the phosphorylation of LRP6 by promoting the interaction between Krt5 and LRP6 of AEC2s, thus activating the Wnt signaling pathway, which is the molecular mechanism of CTGF promoting the proliferation of AEC2s subpopulation. CONCLUSIONS: Our study verifies that CTGF promotes the repair and regeneration of alveoli after acute lung injury by promoting the proliferation of AEC2s subpopulation.

A first-in-human phase I study of SHR-1906, a humanized monoclonal antibody against connective tissue growth factor, in healthy participants.

New therapeutic targets and drugs are urgently needed to halt the fibrosing process in idiopathic pulmonary fibrosis (IPF). SHR-1906 is a novel fully humanized monoclonal antibody against the connective tissue growth factor, which plays an essential role in the genesis of IPF. We assessed the safety, tolerability, pharmacokinetics (PKs), and immunogenicity of single dose SHR-1906 in healthy participants. This was a randomized, double-blind, placebo-controlled, dose-escalation, phase I study. Twelve healthy participants for each dose level were enrolled to receive single ascending doses of SHR-1906 intravenously (1.5, 6, 12, 20, 30, and 45 mg/kg) or placebo and followed for 71 days. The primary end points were safety and tolerability. Treatment-related treatment-emergent adverse events occurred in 25 participants (46.3%) in the SHR-1906 group and 11 (61.1%) in the placebo group. No serious adverse events occurred. Over the dose range investigated, the geometric mean clearance was 0.14-0.63 mL/h/kg, the geometric mean volume of distribution at steady-state was 47.4-75.5 mL/kg, and the terminal elimination half-life was 51.9-349 h. SHR-1906 showed nonlinear PKs. The peak concentration increased in a dose-proportional manner, whereas the area under the concentration-time curve showed a greater than dose-proportional increase. Anti-drug antibodies of SHR-1906 were detected in nine of 54 participants (16.7%). A single dose of SHR-1906 up to 45 mg/kg demonstrated a favorable tolerability profile in healthy participants. The PKs and immunogenicity of SHR-1906 were evaluated, supporting further clinical development.

Context-dependent effects of CCN2 on ß-cell mass expansion and indicators of cell stress in the setting of acute and chronic stress.

Stimulation of functional ß-cell mass expansion can be beneficial for the treatment of type 2 diabetes. Our group has previously demonstrated that the matricellular protein CCN2 can induce ß-cell mass expansion during embryogenesis, and postnatally during pregnancy and after 50% ß-cell injury. The mechanism by which CCN2 stimulates ß-cell mass expansion is unknown. However, CCN2 does not induce ß-cell proliferation in the setting of euglycemic and optimal functional ß-cell mass. We thus hypothesized that ß-cell stress is required for responsiveness to CCN2 treatment. In this study, a doxycycline-inducible ß-cell-specific CCN2 transgenic mouse model was utilized to evaluate the effects of CCN2 on ß-cell stress in the setting of acute (thapsigargin treatment ex vivo) or chronic [high-fat diet or leptin receptor haploinsufficiency (db/+) in vivo] cellular stress. CCN2 induction during 1 wk or 10 wk of high-fat diet or in db/+ mice had no effect on markers of ß-cell stress. However, CCN2 induction did result in a significant increase in ß-cell mass over high-fat diet alone when animals were fed high-fat diet for 10 wk, a duration known to induce insulin resistance. CCN2 induction in isolated islets treated with thapsigargin ex vivo resulted in upregulation of the gene encoding the Nrf2 transcription factor, a master regulator of antioxidant genes, suggesting that CCN2 further activates this pathway in the presence of cell stress. These studies indicate that the potential of CCN2 to induce ß-cell mass expansion is context-dependent and that the presence of ß-cell stress does not ensure ß-cell proliferation in response to CCN2.NEW & NOTEWORTHY CCN2 promotes ß-cell mass expansion in settings of suboptimal ß-cell mass. Here, we demonstrate that the ability of CCN2 to induce ß-cell mass expansion in the setting of ß-cell stress is context-dependent. Our results suggest that ß-cell stress is necessary but insufficient for CCN2 to increase ß-cell proliferation and mass. Furthermore, we found that CCN2 promotes upregulation of a key antioxidant transcription factor, suggesting that modulation of ß-cell oxidative stress contributes to the actions of CCN2.

Key genes and potential drugs in age-related hearing loss: transcriptome analysis of cochlear hair cells in old mice.

This study aimed to dig new molecular mechanisms and medications for age-related hearing loss (ARHL or presbycusis) by extracting common results of publicly available datasets. Based on five datasets (GSE153882, GSE121856, GSE98070, GSE45026, and GSE98071) in studies of cochlear hair cells, we explored the interrelationships among presbycusis-related genes, including gene interactions, enrichment analysis, miRNA-mRNA matching pairs, and potential new drugs. Together, there were 25 common increased mRNAs. A total of 183 drugs can simultaneously target 11 of these mRNAs. In the interaction network, hub genes included: Cbln1, Prl, Mpp6 and Gh. Meanwhile, there were 74 common decreased mRNAs. The hub genes include Cdkn1a, Egr1, and Ctgf. After de-duplication, the 25 common increased mRNAs had 946 matched miRNAs, with 34 decreased ones; and the 74 decreased mRNAs had 1164 matched miRNAs, with 26 increased ones. Between the inhibitors of increased mRNAs and enhancers of decreased mRNAs, there were 26 common drugs. Besides, we discovered six key genes that may play a crucial role in the onset of presbycusis. In conclusion, by jointly analyzing multiple datasets, we found 25 common increased mRNAs (e.g., Cbln1, Prl, Mpp6 and Gh) and 74 common decreased mRNAs (Cdkn1a, Egr1, and Ctgf), as well as 34 potential therapeutic miRNAs and 26 pathogenic miRNAs, and three candidate drugs (calcitriol, diclofenac, and diethylstilbestrol). They may provide new targets and strategies for mechanistic and therapeutic studies in ARHL.

Increased CTGF expression in alveolar epithelial cells by cyclic mechanical stretch: Its mechanism and the therapeutic effect of pirfenidone.

The mechanisms of fibrosis onset and development remain to be elucidated. However, it has been reported that mechanical stretch promotes fibrosis in various organs and cells, and may be involved in the pathogenesis of pulmonary fibrosis. We demonstrated that ventilator-induced lung hyperextension stimulation in mice increased the expression of connective tissue growth factor (CTGF), a profibrotic cytokine, in lung tissue. Increased CTGF expression induced by cyclic mechanical stretch (CMS) was also observed in vitro using A549 human alveolar epithelial cells. Pathway analysis revealed that the induction of CTGF expression by CMS involved MEK phosphorylation. Furthermore, early growth response 1 (Egr-1) was identified as a transcription factor associated with CTGF expression. Finally, the antifibrotic drug pirfenidone significantly reduced CTGF expression, MEK phosphorylation, and Egr-1 levels induced by CMS. Thus, our results demonstrated that profibrotic cytokine CTGF induced by CMS may be a therapeutic target of pirfenidone.

Discovery and Design of Novel Cyclic Peptides as Specific Inhibitors Targeting CCN2 and Disrupting CCN2/EGFR Interaction for Kidney Fibrosis Treatment.

Kidney fibrosis is a serious consequence of chronic kidney disease (CKD), and currently, there is no effective pharmacological treatment available. Cellular communication network-2 (CCN2/CTGF) is an extracellular matrix (ECM) protein that regulates the fibrotic process by activating the epidermal growth factor receptor (EGFR) signaling pathway. We herein present the discovery and structure-activity relationship study of novel peptides targeting CCN2 to develop potent and stable specific inhibitors of the CCN2/EGFR interaction. Remarkably, the 7-mer cyclic peptide OK2 exhibited potent activities to inhibit CCN2/EGFR-induced STAT3 phosphorylation and cellular ECM protein synthesis. Subsequent in vivo studies demonstrated that OK2 significantly alleviated renal fibrosis in a unilateral ureteral obstruction (UUO) mouse model. Moreover, this study first revealed that the peptide candidate could efficiently block CCN2/EGFR interaction through binding to the CT domain of CCN2, providing a new alternative strategy for peptide-based targeting of CCN2 and modulating CCN2/EGFR-mediated biological functions in kidney fibrosis.