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Intervertebral discs are important for maintaining mobility and offer support to the body trunk. If these discs lose their biomechanical features, lower back pain can occur. We previously reported that hepatocyte growth factor (HGF) promotes cell proliferation and suppresses apoptosis, inflammation, and matrix degradation in nucleus pulposus (NP) cells. In the present study, we investigated the molecular mechanisms of how HGF promotes the proliferation of NP cells in hypoxic conditions. Hypoxic stimulation promoted modest cell proliferation, which was further upregulated by HGF. Expression of hypoxia-inducible factor (HIF-1α) protein, which contributes to the maintenance of homeostasis in NP cells, was also upregulated in hypoxia-treated cell groups; HGF further increased HIF-1α expression in NP cells. Additionally, knockdown of HIF-1α expression significantly reduced the proliferation of NP cells. An MAPK inhibitor inhibited the expression of HIF-1α and pERK, as well as cell proliferation in a dose-dependent manner. Similarly, inhibiting the PI3K/Akt and STAT3 pathways also decreased the expression of HIF-1α and cell proliferation. These results show that under hypoxic conditions, HGF promotes NP cell proliferation via HIF-1α-, MAPK-, PI3K/Akt-, and STAT3-mediated signaling which is involved in this pathway. The control of these signaling pathways may be a target for potential therapeutic strategies for the treatment of disc degeneration in hypoxic conditions., (© 2020 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Objective: To select a suitable combination of classic angiogenic and vascular stabilization factors to improve the proliferation and maturity of neovascularization of lung tissue in a rat pulmonary arterial hypertension (PAH) model., Methods: PAH rat model was established by intraperitoneal injection of monocrotaline. Proangiogenic factors hepatocyte growth factor (HGF) and vascular endothelial growth factor (VEGF), as well as vascular stabilization factors angiopoietin-1 (Ang-1), platelet-derived growth factor, and transforming growth factor-beta were transfected by pairs into the lung tissue of rats with PAH through lentivirus. Four weeks later, pulmonary artery angiography and hemodynamic parameters were determined to testify the remission of PAH. Immunofluorescence staining and Western blot were performed to investigate the structure and function of neovascularization., Results: The pulmonary artery pressure and weight index of the right ventricle in HGF+Ang-1 and VEGF+Ang-1 groups were significantly decreased compared with vehicle group. The contrast medium filling time and right pulmonary artery root diameter were also significantly decreased. In addition, the maturity and perfusion of neovascularization in HGF+Ang-1 and VEGF+Ang-1 groups were promoted compared to vehicle group, and vascular leakage was reduced. Finally, the adherens junction integrity of vascular endothelial cells in HGF+Ang-1 and VEGF+Ang-1 combinations was upregulated compared with other combinations., Conclusions: HGF+Ang-1 transfection and VEGF+Ang-1 transfection alleviate PAH by promoting maturation and stability of new blood vessels, which may be potential candidates for PAH treatment., (Copyright © 2020 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.)

Incretins, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), are hormones secreted from small intestine accompanied with oral intake. We previously showed that transforming growth factor (TGF)-α stimulates the migration of hepatocellular carcinoma (HCC) cells via mitogen-activated protein (MAP) kinases, AKT and Rho-kinase. However, it remains to be elucidated whether incretins affect HCC cell functions. In the present study, therefore, we investigated whether incretins affect the migration of HCC cells using human HCC-derived HuH7 cells. GLP-1, but not GIP, reduced both TGF-α- and hepatocyte growth factor (HGF)-induced cell migration. IBMX, an inhibitor of cyclic nucleotide phosphodiesterase, enhanced the suppressive effect of GLP-1. GLP-1 attenuated the phosphorylation of stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) by TGF-α and HGF. Our results strongly suggest that GLP-1 suppresses TGF-α- and HGF-induced migration of HCC cells through inhibiting the SAPK/JNK signaling pathway, and that the inhibition by GLP-1 is due to cAMP production., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Rationale: Epithelial-mesenchymal transition (EMT) has been recognized as an important step toward high invasion and metastasis of many cancers including hepatocellular carcinoma (HCC), while the mechanism for EMT promotion is still ambiguous. Methods: The dynamic alterations of site-specific glycosylation during HGF/TGF-β1-induced EMT process of three HCC cell lines were systematically investigated using precision glycoproteomic methods. The possible roles of EMT-related glycoproteins and site-specific glycans were further confirmed by various molecular biological approaches. Results: Using mass spectrometry-based glycoproteomic methods, we totally identified 2306 unique intact glycopeptides from SMMC-7721 and HepG2 cell lines, and found that core-fucosylated glycans were accounted for the largest proportion of complex N -glycans. Through quantification analysis of intact glycopeptides, we found that the majority of core-fucosylated intact glycopeptides from folate receptor α (FOLR1) were up-regulated in the three HGF-treated cell lines. Similarly, core-fucosylation of FOLR1 were up-regulated in SMMC-7721 and Hep3B cells with TGF-β1 treatment. Using molecular approaches, we further demonstrated that FUT8 was a driver for HGF/TGF-β1-induced EMT. The silencing of FUT8 reduced core-fucosylation and partially blocked the progress of HGF-induced EMT. Finally, we confirmed that the level of core-fucosylation on FOLR1 especially at the glycosite Asn-201 positively regulated the cellular uptake capacity of folates, and enhanced uptake of folates could promote the EMT of HCC cells. Conclusions: Based on the results, we proposed a potential pathway for HGF or TGF-β1-induced EMT of HCC cells: HGF or TGF-β1 treatment of HCC cells can increase the expression of glycosyltransferase FUT8 to up-regulate the core-fucosylation of N -glycans on glycoproteins including the FOLR1; core-fucosylation on FOLR1 can then enhance the folate uptake capacity to finally promote the EMT progress of HCC cells., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Hepatocyte growth factor (HGF) promotes the neurite outgrowth of sensory neurons in developmental stages, but its role in injured peripheral nerves in adult mice remains largely been unexplored. In this study, we investigated the role of HGF in the regeneration of injured peripheral nerves using cultured dorsal root ganglions (DRGs). When cells were treated with HGF protein, the length of the neurite was increased 1.4-fold compared to the untreated control group. HGF greatly increased the level of phosphorylated STAT3 at serine 727 [pSTAT3 (Ser 727)], thereby translocating the protein to the mitochondria. HGF treatment increased the activity of mitochondrial complex I. When DRGs were cultured in the presence of U0126, a pharmacological inhibitor of Erk, the HGF-mediated increase in neurite outgrowth and the level of pSTAT3 (Ser 727) were both suppressed. Taken together, these results suggest that the HGF/c-met pathway might promote neurite outgrowth by controlling mitochondrial activity through the HGF/Erk/STAT3 axis., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Induction of intrinsic liver regeneration is an unmet need that can be achieved by temporally activating key hepatocyte regenerative pathways. Here, we establish an efficient, safe, non-integrative method to transiently express hepatocyte-growth-factor (HGF) and epidermal-growth-factor (EGF) in hepatocytes via nucleoside-modified, lipid-nanoparticle-encapsulated mRNA (mRNA-LNP) delivery in mice. We confirm specific hepatotropism of mRNA-LNP via intravenous injection of firefly luciferase encoding mRNA-LNP, with protein expression lasting about 3 days. In the liver, virtually all hepatocytes are transfected along with a subpopulation of endothelial and Kupffer cells. In homeostasis, HGF mRNA-LNP efficiently induce hepatocyte proliferation. In a chronic liver injury mouse model recapitulating non-alcoholic fatty liver disease, injections of both HGF and EGF mRNA-LNP sharply reverse steatosis and accelerate restoration of liver function. Likewise, HGF and EGF mRNA-LNP accelerate liver regeneration after acetaminophen-induced acute liver injury with rapid return to baseline ALT levels. This study introduces mRNA-LNP as a potentially translatable safe therapeutic intervention to harness liver regeneration via controlled expression of endogenous mitogens in vivo.

ADAMTSs (A Disintegrin And Metalloproteinase with ThromboSpondin motifs) are secreted proteases dependent on Zn 2+ /Ca 2+ , involved in physiological and pathological processes and are part of the extracellular matrix (ECM). Here, we investigated if ADAMTS-1 is required for invasion and migration of cells and the possible mechanism involved. In order to test ADAMTS-1's role in ovarian cancer cells (CHO, NIH-OVCAR-3 and ES2) and NIH-3 T3 fibroblasts, we modified the levels of ADAMTS-1 and compared those to parental. Cells exposed to ADAMTS-1-enriched medium exhibited a decline in cell migration and invasion when compared to controls with or without a functional metalloproteinase domain. The opposite was observed in cells when ADAMTS-1 was deleted via the CRISPR/Cas9 approach. The decline in ADAMTS-1 levels enhanced the phosphorylated form of Src and FAK. We also evaluated the activities of cellular Rho GTPases from cell lysates using the GLISA® kit. The Cdc42-GTP signal was significantly increased in the CRISPR ADAMTS-1 ES-2 cells. By a Förster resonance energy transfer (FRET) biosensor for Cdc42 activity in ES-2 cells we demonstrated that Cdc42 activity was strongly polarized at the leading edge of migrating cells with ADAMTS-1 deletion, compared to the wild type cells. As conclusion, ADAMTS-1 inhibits proliferation, polarization and migration., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Imbalance between free radicals and antioxidants causes oxidative stress by the accumulation of reactive oxygen species (ROS) in the tissues and organs. Oxidative stress occurs in many damaged conditions, and the increase of ROS and reduction of antioxidants enhances inflammation, apoptosis, fibrosis and may worsen the pathology leading to organ failure. The potential therapies aim to increase antioxidants and decrease ROS. Mesenchymal stem cells (MSCs) isolated from the stroma of various tissues are multipotent cells and have beneficial effects on several diseases with their immunomodulation and regeneration capacities. MSCs trigger the proliferation of the cells with various secretory factors, reduce oxidative stress and decrease apoptosis, inflammation, fibrosis and thus, increase regeneration. However, survival, engraftment, and differentiation problems of transplanted MSCs restrict their protective and regenerative effects. Preconditioning of MSCs with several factors, such as cytokines, hypoxia, chemical agents, pharmacological drugs, physical factors and growth factors, enhances their repairing efficacy for injury and disease models. This review is mainly focused on insulin-like growth factor (IGF-1) and hepatocyte growth factor (HGF), and discusses the research on MSC priming/induction with IGF-1 and HGF stimulation either by supplementation or overexpression that can enhance the regenerative potential of MSCs on various oxidative stress conditions such as acute/chronic kidney diseases, lung injury, cancer, metabolic and cardiovascular diseases., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Diabetic retinopathy (DR) is a disease that causes blindness due to vascular leakage or abnormal angiogenesis. Hepatocyte growth factor (HGF) is increased in the serum or vitreous fluid in proliferative diabetic retinopathy (PDR) patients, although the effect of HGF on the blood vessels remains unclear. This study focused on the effect of HGF on pericyte (PC) survival and endothelial cell (EC) permeability. It was demonstrated that HGF was increased in the diabetic mouse retina. However, HGF prevented PC apoptosis caused by TNF-α, which increased in the diabetic retinas both in vitro and in vivo. In addition, HGF was involved in PC survival by increasing the Akt signaling pathway. Moreover, HGF strengthened the EC tight junction in co-cultures of PCs and ECs by promoting PC survival, thereby reducing EC permeability. These results suggest that HGF may play a role in the prevention of increased vascular leakage by inhibiting the PC loss that occurs in DR to some extent. However, careful HGF reduction in DR might avoid an increase in PC loss., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: Pancreatic stellate cells (PSCs) play key roles in the pancreatic tumor microenvironment and are considered to contribute to chemoresistance. PSCs can participate in malignant behaviors of pancreatic carcinoma (PC) by secreting hepatocyte growth factor (HGF). The objective of this research was to explore the potential molecular mechanism of HGF on gemcitabine (GEM) chemoresistance of PC., Materials and Methods: HGF, c-Met, E-Cadherin and Vimentin levels were examined by quantitative real-time polymerase chain reaction (qRT-PCR). The changes of HGF level were detected by ELISA. The half maximal inhibitory concentration, the growth inhibitions and apoptosis of pancreatic cancer cells (PCCs) were respectively assayed using CCK-8 and flow cytometry. Associated proteins were measured using western blot and cell immunofluorescence assay., Key Findings: PSCs strongly expressed HGF, and its receptor c-Met was expressed in PCCs. PCCs exerted a positive regulative effect on HGF production. HGF neutralizing antibody AMG102 could effectively reduce the HGF level in PSC-conditioned medium (PSC-CM). PSC-CM promoted chemoresistance in PCCs. When exposed to PSC-CM, PCCs underwent epithelial-to-mesenchymal transition (EMT), and c-Met was also activated. Recombinant human HGF had the same protective effect. Blocking the HGF/c-Met axis with a c-Met inhibitor PHA665752 and AMG102 reduced the phosphorylation level of c-Met (p-c-Met) and attenuated EMT and chemoresistance. P-c-Met overexpression resulted in activation of the PI3K/Akt pathway, and inhibition of PI3K/Akt signaling with LY294002 reversed chemoresistance and EMT., Significance: PSCs can activate the c-Met/PI3K/Akt pathway in PCCs via paracrine HGF, induce EMT of PCCs and inhibit cancer cell apoptosis, thus enhance chemoresistance to Gem in PCCs., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background and Aims: Ulcerative colitis [UC] is associated with excessive neutrophil infiltration and collateral tissue damage, but the link is not yet completely understood. Since c-MET receptor tyrosine kinase [MET] is required for neutrophil chemoattraction and cytotoxicity in response to its ligand hepatocyte growth factor [HGF], we aimed to identify the function of HGF-MET signalling in neutrophils in UC patients and in mice during intestinal inflammation., Methods: Serum and colonic biopsies from healthy controls and UC patients with active [Mayo endoscopic subscore 2-3] and inactive [Mayo endoscopic subscore 0-1] disease were collected to assess the level of serum and colonic HGF. Disease progression and immune cell infiltration were assessed during dextran sodium sulphate [DSS] colitis in wild-type and MRP8-Cre MET-LoxP mice., Results: Increased mucosal HGF expression was detected in patients with active UC, and in mice during the inflammatory phase of DSS colitis. Similarly, serum HGF was significantly increased in active UC patients and positively correlated with C-reactive protein and blood neutrophil counts. Flow cytometric analysis also demonstrated an upregulation of colonic MET+ neutrophils during DSS colitis. Genetic ablation of MET in neutrophils reduced the severity of DSS-induced colitis. Concomitantly, there was a decreased number of TH17 cells, which could be due to a decreased production of IL-1β by MET-deficient neutrophils., Conclusions: These data highlight the central role of neutrophilic HGF-MET signalling in exacerbating damage during intestinal inflammation. Hence, selective blockade of this pathway in neutrophils could be considered as a novel therapeutic approach in UC., (© The Author(s) 2020. Published by Oxford University Press on behalf of European Crohn’s and Colitis Organisation. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Head and neck squamous cell carcinoma (HNSCC) is a widespread disease with a low survival rate and a high risk of recurrence. Nowadays, immune checkpoint inhibitor (ICI) treatment is approved for HNSCC as a first-line treatment in recurrent and metastatic disease. ICI treatment yields a clear survival benefit, but overall response rates are still unsatisfactory. As shown in different cancer models, hepatocyte growth factor/mesenchymal-epithelial transition (HGF/Met) signaling contributes to an immunosuppressive microenvironment. Therefore, we investigated the relationship between HGF and programmed cell death protein 1 (PD-L1) expression in HNSCC cell lines. The preclinical data show a robust PD-L1 induction upon HGF stimulation. Further analysis revealed that the HGF-mediated upregulation of PD-L1 is MAP kinase-dependent. We then hypothesized that serum levels of HGF and soluble programmed cell death protein 1 (sPD-L1) could be potential markers of ICI treatment failure. Thus, we determined serum levels of these proteins in 20 HNSCC patients before ICI treatment and correlated them with treatment outcomes. Importantly, the clinical data showed a positive correlation of both serum proteins (HGF and sPD-L1) in HNSCC patient's sera. Moreover, the serum concentration of sPD-L1 was significantly higher in ICI non-responsive patients. Our findings indicate a potential role for sPD-L1 as a prognostic marker for ICI treatment in HNSCC.

Multiple myeloma (MM)-induced bone disease occurs through hyperactivation of osteoclasts by several factors secreted by MM cells. MM cell-secreted factors induce osteoclast differentiation and activation via direct and indirect actions including enhanced expression of receptor activator of nuclear factor κB ligand (RANKL) in osteoblasts and bone marrow stromal cells (BMSCs). Hepatocyte growth factor (HGF) is elevated in MM patients and is associated with MM-induced bone disease, although the mechanism by which HGF promotes bone disease remains unclear. In the present study, we demonstrated that HGF induces RANKL expression in osteoblasts and BMSCs, and investigated the mechanism of induction. We found that HGF and MM cell supernatants induced RANKL expression in ST2 cells, MC3T3-E1 cells, and mouse BMSCs. In addition, HGF increased phosphorylation of Met and nuclear factor κB (NF-κB) in ST2 cells, MC3T3-E1 cells, or mouse BMSCs. Moreover, Met and NF-κB inhibitors suppressed HGF-induced RANKL expression in ST2 cells, MC3T3-E1 cells, and mouse BMSCs. These results indicated that HGF promotes RANKL expression in osteoblasts and BMSCs via the Met/NF-κB signaling pathway, and Met and NF-κB inhibitors suppressed HGF-induced RANKL expression. Our findings suggest that Met and NF-κB inhibitors are potentially useful in mitigating MM-induced bone disease in patients expressing high levels of HGF.

Angiotensin II (Ang II) contributes to renal dysfunction, while hepatocyte growth factor (HGF) protects against renal dysfunction. However, the relationship between Ang II and HGF in chronic kidney disease (CKD) remains unknown. This study aimed to investigate the effect of HGF on Ang II expression in CKD. A rat model of CKD was established using female Wistar rats subjected to 5/6 nephrectomy (5/6 Nx). HGF was overexpressed in rat renal tissues using PCI-neo-HGF. Immunohistochemical staining and western blot analysis of renal Ang II expression were performed in 5/6 Nx rats treated with vehicle (negative control), Lotensin (positive control), or HGF for different periods of time (before 5/6 Nx, 5 and 9 weeks after 5/6 Nx). Compared with the 0-week group (before 5/6 Nx group), the protein expression of Ang II was significantly induced in rat renal tissues at 5 and 9 weeks after 5/6 Nx ( p <0.05), suggesting the possible involvement of Ang II in 5/6 Nx-induced CKD. Importantly, HGF treatment for 5 or 9 weeks markedly inhibited renal Ang II expression and greatly improved the renal morphology in 5/6 Nx rats, compared with the vehicle-treated group ( p <0.05). The effects of HGF on renal Ang II expression and renal morphology were similar to those of Lotensin, suggesting that HGF may protect against 5/6 Nx-induced CKD through downregulating Ang II. HGF is a novel regulator of Ang II expression and plays a protective role in 5/6 Nx-induced CKD., (© 2020 by the Association of Clinical Scientists, Inc.)

Background: Mesenchymal stem cells (MSCs) have a limited self-renewal ability, impaired multi-differentiation potential, and undetermined cell senescence during in vitro series expansion. To address this concern, we investigated the effects of the microenvironment provided by stem cells from human exfoliated deciduous teeth (SHED) in maintaining the stemness of human bone marrow mesenchymal stem cells (hBMSCs) and identified the key factors and possible mechanisms responsible for maintaining the stemness of MSCs during long-term expansion in vitro., Methods: The passage 3 (P3) to passage 8 (P8) hBMSCs were cultured in the conditioned medium from SHED (SHED-CM). The percentage of senescent cells was evaluated by β-galactosidase staining. In addition, the osteogenic differentiation potential was analyzed by reverse transcription quantitative PCR (RT-qPCR), Western blot, alizarin red, and alkaline phosphatase (ALP) staining. Furthermore, RT-qPCR results identified hepatocyte growth factor (HGF) and stem cell factor (SCF) as key factors. Thus, the effects of HGF and SCF on mitochondrial function were assessed by measuring the ROS and mitochondrial membrane potential levels. Finally, selected mitochondrial-related proteins associated with the PI3K/AKT, ERK1/2, and STAT3 signaling pathways were investigated to determine the effects of HGF and SCF in preserving the mitochondrial function of hBMSCs during long-term expansion., Results: SHED-CM had significantly enhanced the cell proliferation, reduced the senescent cells, and maintained the osteogenesis and pro-angiogenic capacity in P8 hBMSCs during long-term expansion. In addition, hBMSCs treated with 100 ng/ml HGF and 10 ng/ml SCF had reduced ROS levels and preserved mitochondrial membrane potential compared with P8 hBMSCs during long-term expansion. Furthermore, HGF and SCF upregulated the expression of mitochondrial-related proteins associated with the PI3K/AKT, ERK1/2, and STAT3 signaling pathways, possibly contributing to the maintenance of hBMSCs stemness by preserving mitochondrial function., Conclusion: Both HGF and SCF are key factors in maintaining the stemness of hBMSCs by preserving mitochondrial function through the expression of proteins associated with the PI3K/AKT, ERK1/2, and STAT3 signaling pathways. This study provides new insights into the anti-senescence capability of HGF and SCF, as well as new evidence for their potential application in optimizing the long-term culture of MSCs.

Acute myocardial infarction (AMI) is recognized to be a severe threat to people's health conditions and life quality. The accumulation of hepatocyte growth factor (HGF) in ischemic myocardium has been observed in both processes of experimental ischemia and reperfusion (I/R) and permanent coronary artery occlusion. The aim of the study was to investigate the effect of HGF on myocardial cell apoptosis, ventricular remodeling and cardiac function after myocardial infarction (MI) in diabetic rats, and to explore whether the effect is mediated by HGF/c-Met signaling pathway. MI significantly increases LVWI and RVWI and myocardial apoptotic index, and up-regulates the expression of HGF and c-Met at mRNA and protein levels in MI control group. The LVWI and RVWI, and myocardial apoptosis were reduced by treatment with HGF, which also increased the myocardial cell viability and the expression of HGF and c-Met. In summary, HGF significantly attenuates myocardial apoptosis and improves cardiac function after AMI in diabetic rats by further enhancing the activation of HGF/c-Met pathway.

Previous studies showed that the hepatocyte growth factor (HGF)-Met receptor axis plays long-lasting cardioprotection against doxorubicin anti-cancer therapy. Here, we explored the mechanism(s) underlying the HGF protective effect. DNA damage was monitored by histone H2AX phosphorylation and apoptosis by proteolytic cleavage of caspase 3. In doxorubicin-treated H9c2 cardiomyoblasts, the long-lasting cardioprotection is mediated by activation of the Ras/Raf/Mek/Erk (extracellular signal-regulated kinase 1,2) signaling pathway and requires Stat3 (signal transducer and activator of transcription 3) activation. The HGF protection was abrogated by the Erk1,2 inhibitor, PD98059. This translated into reduced Y705 phosphorylation and impaired nuclear translocation of Stat3, showing crosstalk between Erk1,2 and Stat3 signaling. An array of 29 cytokines, known to activate Stat3, was interrogated to identify the molecule(s) linking the two pathways. The analysis showed a selective increase in expression of the tissue inhibitor of metalloproteinases-1 (Timp1). Consistently, inhibition in cardiomyoblasts of Timp1 translation by siRNAs blunted both Stat3 activation and the cardioprotective effect of HGF. Thus, Timp1 is responsible for the generation of a feed-forward loop of Stat3 activation and helps cardiomyocytes to survive during the genotoxic stress induced by anthracyclines.

Posterior capsule opacification (PCO) as a result of proliferation and fibrogenesis of lens epithelial cells (LECs) is the most frequent long‑term complication of modern cataract surgery. LECs may undergo epithelial‑mesenchymal transition (EMT) that resembles the morphological and molecular characteristics of PCO. A pre‑identified novel, hepatocyte growth factor (HGF)‑derived peptide H‑RN, was reported to exhibit anti‑angiogenic activity and anti‑inflammatory effects in ocular cells both in vitro and in vivo. However, the role of H‑RN in the promotion of the development of EMT in LECs is unknown. In the present study, the effects of H‑RN on the development of EMT induced by transforming growth factor (TGF)‑β in human LECs, and the possible signaling pathways participating in this process were investigated. The results showed that H‑RN promoted the expression of the EMT‑associated markers, α‑smooth muscle actin and fibronectin, whereas the expression of E‑cadherin and connexin 43 were reduced. The morphological changes typically associated with EMT seen in LECs induced by TGF‑β2 were inhibited by H‑RN, which was consistent with the effects of a TGF‑β2 inhibitor, SB431542. Smad2 and Smad3 phosphorylation induced by TGF‑β2 were reduced by H‑RN, and phosphorylation of Akt, mTOR and P70S6K induced by TGF‑β2 were also notably reduced by H‑RN in LECs. Therefore, the results of the present study showed that H‑RN treatment significantly suppressed the development of EMT induced by TGF‑β2, at least partially through the TGF‑β/Smad and Akt/mTOR signaling pathways in human LECs. The present study highlights that H‑RN, a novel HGF‑derived peptide, may be a novel therapeutic agent for prevention and treatment of PCO.

In the light of hepatocyte growth factor (HGF) the inhibiting role on the expression of hepcidin, we hypothesized that HGF might be able to reduce cell and tissue iron by increasing ferroportin 1 (Fpn1) content and Fpn1-mediated iron release from cells and tissues. The hypothesized ability of HGF to reduce iron might be one of the mechanisms associated with its neuroprotective action under the conditions of ischemia/reperfusion (I/R). Here, we investigated the effects of HGF on the expression of hepcidin as well as transferrin receptor 1 (TfR1), divalent metal transporter 1 (DMT1), Fpn1, ferritin and iron regulatory proteins (IRPs) in oxygen-glucose deprivation and reoxygenation (OGD/R)-treated PC12 cells by real-time PCR and Western blot analysis. We demonstrated that HGF could completely reverse the OGD/R-induced reduction in Fpn1 and IRP1 expression and increase in ferritin light chain protein and hepcidin mRNA levels in PC12 cells. It was concluded that HGF protects PC12 cells against OGD/R-induced injury mainly by reducing cell iron contents via the up-regulation of Fpn1 and increased Fpn1-mediated iron export from cells. Our findings suggested that HGF may also be able to ameliorate OGD/R or I/R-induced overloading of brain iron by promoting Fpn1 expression., (© 2020 The Author(s).)

Receptor tyrosine kinases (RTKs) orchestrate cell motility and differentiation. Deregulated RTKs may promote cancer and are prime targets for specific inhibitors. Increasing evidence indicates that resistance to inhibitor treatment involves receptor cross-interactions circumventing inhibition of one RTK by activating alternative signaling pathways. Here, we used single-molecule super-resolution microscopy to simultaneously visualize single MET and epidermal growth factor receptor (EGFR) clusters in two cancer cell lines, HeLa and BT-20, in fixed and living cells. We found heteromeric receptor clusters of EGFR and MET in both cell types, promoted by ligand activation. Single-protein tracking experiments in living cells revealed that both MET and EGFR respond to their cognate as well as non-cognate ligands by slower diffusion. In summary, for the first time, we present static as well as dynamic evidence of the presence of heteromeric clusters of MET and EGFR on the cell membrane that correlates with the relative surface expression levels of the two receptors.

The current treatment for liver failure is restricted to surgical liver transplantation, which is technically complicated, limited by the shortage of available organs and presents major risks to the patient. Bone marrow mesenchymal stem cells (BMSCs) represent promising sources of hepatocyte-like cells for cell transplantation treatment. However, a safe and efficient induction method for their differentiation remains to be defined. Here we further optimized an effective technique by combining high-dose treatment with hepatocyte growth factor (HGF) and ultrasound stimulation. The optimized ultrasound parameter (1.0 W/cm 2 intensity, 1 MHz frequency, 20% duty cycle, 100 Hz pulse repetition frequency, 60-s irradiation duration, triple times in three days) combined with different HGF doses (10, 20 and 50 ng/ml) was used to treat BMSCs. The results showed that the specific hepatic markers, including α-fetoprotein (αFP/AFP), cytokeratin 18 (CK18), albumin (ALB) and glycogen, were increased in a dose-dependent manner. Their concentration was then further increased when ultrasound irradiation was administered ( P < 0.05), as indicated by PCR, Western blot and immunofluorescence staining as well as a glycogen synthesis test. Furthermore, analysis of the hepatocyte-derived chemokines showed elevated stromal cell-derived factor 1alpha (SDF-1α) and C-X-C chemokine receptor type 4 (CXCR4) after HGF treatment. Again, concentrations of those chemokines were further increased by ultrasound radiation ( P < 0.05). The observed increased effect was sustained for 21 days. To summarize, we further defined the optimal combination of HGF and ultrasound treatment to increase the differentiation and chemotaxis of BMSCs in a safe, sustained and efficient manner. These findings provide a new perspective for stem cell orientation in the field of tissue engineering.

Cholestasis is a clinical syndrome common to a large number of hepatopathies, in which either bile production or its transit through the biliary tract is impaired due to functional or obstructive causes; the consequent intracellular retention of toxic biliary constituents generates parenchyma damage, largely via oxidative stress-mediated mechanisms. Hepatocyte growth factor (HGF) and its receptor c-Met represent one of the main systems for liver repair damage and defense against hepatotoxic factors, leading to an antioxidant and repair response. In this study, we evaluated the capability of HGF to counteract the damage caused by the model cholestatic agent, α-naphthyl isothiocyanate (ANIT). HGF had clear anti-cholestatic effects, as apparent from the improvement in both bile flow and liver function test. Histology examination revealed a significant reduction of injured areas. HGF also preserved the tight-junctional structure. These anticholestatic effects were associated with the induction of basolateral efflux ABC transporters, which facilitates extrusion of toxic biliary compounds and its further alternative depuration via urine. The biliary epithelium seems to have been also preserved, as suggested by normalization in serum GGT levels, CFTR expression and cholangyocyte primary cilium structure our results clearly show for the first time that HGF protects the liver from a cholestatic injury., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Flavonol, which is found abundantly in plants such as fruits and vegetables, belongs to the family of flavonoid, natural polyphenols. Quercetin, one of the flavonol, reportedly has anti-cancer effects and prevents the proliferation of various cancer cells, including hepatocellular carcinoma (HCC). However, the effects of quercetin on HCC cells migration have not yet been clarified. We have previously shown that the migration of human HCC-derived HuH7 cells induced by hepatocyte growth factor (HGF) or transforming growth factor-α (TGF-α) is mediated through p38 MAPK and AKT. In this study, we investigated whether quercetin affects the HGF- or TGF-α-induced migration of HuH7 cells. Quercetin significantly suppressed both HGF- and TGF-α-induced migration of HuH7 cells in a dose-dependent manner. In addition, myricetin, another flavonol, also showed significant inhibition of the cell migration. Each HGF- and TGF-α-induced autophosphorylation of receptors were not affected by quercetin or myricetin. Quercetin did not suppress HGF- or TGF-α-induced p38 MAPK phosphorylation. On the contrary, quercetin and myricetin inhibited the growth factors-induced phosphorylation of AKT. Our results strongly suggest that quercetin suppresses the growth factor-induced migration of HCC cells by inhibiting the signaling pathway of AKT but not p38 MAPK., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The clinical use of human bone marrow-derived mesenchymal stem cells (BM-MSCs) has been hampered by their poor performance after transplantation into failing hearts. Here, to improve the therapeutic potential of BM-MSCs, we developed a strategy termed in vivo priming in which BM-MSCs are primed in vivo in myocardial infarction (MI)-induced hearts through genetically engineered hepatocyte growth factor-expressing MSCs (HGF-eMSCs) that are encapsulated within an epicardially implanted 3D cardiac patch. Primed BM-MSCs through HGF-eMSCs exhibited improved vasculogenic potential and cell viability, which ultimately enhanced vascular regeneration and restored cardiac function to the MI hearts. Histological analyses further demonstrated that the primed BM-MSCs survived longer within a cardiac patch and conferred cardioprotection evidenced by substantially higher numbers of viable cardiomyocytes in the MI hearts. These results provide compelling evidence that this in vivo priming strategy can be an effective means to enhance the cardiac repair of MI hearts., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Evodiamine (EVO) is an indoloquinazoline alkaloid that exerts its various anti-oncogenic actions by blocking phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt), mitogen-activated protein kinase (MAPK), c-Met, and nuclear factor kappa B (NF-κB) signaling pathways, thus leading to apoptosis of tumor cells. We investigated the ability of EVO to affect hepatocyte growth factor (HGF)-induced c-Met/Src/STAT3 activation cascades in castration-resistant prostate cancer (CRPC). First, we noted that EVO showed cytotoxicity and anti-proliferation activities in PC-3 and DU145 cells. Next, we found that EVO markedly inhibited HGF-induced c-Met/Src/STAT3 phosphorylation and impaired the nuclear translocation of STAT3 protein. Then, we noted that EVO arrested the cell cycle, caused apoptosis, and downregulated the expression of various carcinogenic markers such as B-cell lymphoma 2 (Bcl-2), B-cell lymphoma-extra large (Bcl-xL), cyclin D1, cyclooxygenase 2 (COX-2), survivin, vascular endothelial growth factor (VEGF), and matrix metallopeptidases 9 (MMP-9). Moreover, it was observed that in cPC-3 and DU145 cells transfected with c-Met small interfering RNA (siRNA), Src/STAT3 activation was also mitigated and led to a decrease in EVO-induced apoptotic cell death. According to our results, EVO can abrogate the activation of the c-Met/Src/STAT3 signaling axis and thus plays a role as a robust suppressor of tumor cell survival, proliferation, and angiogenesis.

Lenvatinib is a long-awaited alternative to sorafenib for the first-line targeted therapy of patients with advanced hepatocellular carcinoma (HCC). However, resistance to lenvatinib has also become a major obstacle to improving the prognosis of HCC patients. The underlying molecular mechanisms contributing to lenvatinib resistance in HCC are largely unknown. HGF/c-MET axis activation is related to tumor progression and several hallmarks of cancer and is considered as the key contributor to drug resistance. In the present study, we focused on the role of the HGF/c-MET axis in mediating lenvatinib resistance in HCC cells. We showed that HGF reduced the antiproliferative, proapoptotic, and anti-invasive effects of lenvatinib on HCC cells with high c-MET expression but did not significantly affect HCC cells with low c-MET expression. The c-MET inhibitor PHA-665752 rescued HCC cells from HGF-induced lenvatinib resistance. Furthermore, HGF/c-MET activated the downstream PI3K/AKT and MAPK/ERK pathways and promoted epithelial-mesenchymal transition (EMT) in HCC cells. Collectively, our results suggested that combining lenvatinib treatment with a c-MET inhibitor may improve its systemic therapeutic efficacy in HCC patients with high c-MET expression.

Tuberculosis (TB) is one of the deadliest infectious diseases in humankind history. Although, drug sensible TB is slowly decreasing, at present the rise of TB cases produced by multidrug-resistant (MDR) and extensively drug-resistant strains is a big challenge. Thus, looking for new therapeutic options against these MDR strains is mandatory. In the present work, we studied, in BALB/c mice infected with MDR strain, the therapeutic effect of supra-pharmacological doses of the conventional primary antibiotics rifampicin and isoniazid (administrated by gavage or intratracheal routes), in combination with recombinant human hepatocyte growth factor (HGF). This high dose of antibiotics administered for 3 months, overcome the resistant threshold of the MDR strain producing a significant reduction of pulmonary bacillary loads but induced liver damage, which was totally prevented by the administration of HGF. To address the long-term efficiency of this combined treatment, groups of animals after 1 month of treatment termination were immunosuppressed by glucocorticoid administration and, after 1 month, mice were euthanized, and the bacillary load was determined in lungs. In comparison with animals treated only with a high dose of antibiotics, animals that received the combined treatment showed significantly lower bacterial burdens. Thus, treatment of MDR-TB with very high doses of primary antibiotics particularly administrated by aerial route can produce a very good therapeutic effect, and its hepatic toxicity can be prevented by the administration of HGF, becoming in a new treatment modality for MDR-TB., (© 2019 Wiley Periodicals, Inc.)

Ischemia-reperfusion (I/R)-induced organ injury is a serious health problem worldwide, and poor recovery of acute phase injury leads to chronic fibrosis and further organ dysfunction. Thus, a more precise approach to enhance tissue repair is needed. By using a renal I/R model, we aimed to evaluate the role of a hydrogel-based dual-drug delivery platform on promoting tissue repair. An injectable, self-assembling peptide/heparin (SAP/Hep) hydrogel was used to co-deliver TNF-α neutralizing antibody (anti-TNF-α) and hepatocyte growth factor (HGF). The microstructure and controlled release properties of KLD2R/Hep hydrogel were analyzed. The effects of the drug-loaded hydrogel (SAP-drug) on renal injury were evaluated in mice with I/R injury. In vitro, the SAP/Hep hydrogel allowed for a faster release of anti-TNF-α with a sustained release of HGF, and both drugs maintained their bioactivities after release. In vivo, combined anti-TNF-α/HGF showed better renal protective potential than anti-TNF-α or HGF alone. SAP-drug (anti-TNF-α/HGF in SAP hydrogel) treatment reduced the level of serum creatinine (Scr), blood urea nitrogen (BUN), tubular apoptosis, renal inflammatory factors, and macrophage infiltration compared to Free-drug (anti-TNF-α/HGF in solution) or SAP alone. Moreover, the SAP-drug group had better efficacy on promoting tubular cell proliferation and dedifferentiation than SAP or Free-drug alone, and thus reduced chronic renal fibrosis in I/R mice. This study highlighted that SAP could sequentially deliver the two drugs to achieve anti-inflammatory and pro-proliferative effects with one injection and thus is a promising delivery platform for tissue repair. STATEMENT OF SIGNIFICANCE: Ischemia-reperfusion (I/R)-induced organ injury is a serious health issue, and delayed tissue repair leads to chronic fibrosis and organ failure. Systemic administration of anti-inflammatory agents or growth factors have shown some benefits on I/R injury, but their therapeutic efficacy was limited by side effects, poor bioavailability, and absent key signals of tissue repair. To address these issues, a hydrogel-based drug co-delivery platform was used to treat I/R injury. This platform could achieve sequential release kinetics with faster rate of anti-TNF-ɑ and slower rate of HGF, and effectively promoted tissue repair by targeting inflammation and proliferation in mice with renal I/R. This nanoscale delivery platform represents a promising strategy for solid organs (heart, liver and kidney) regeneration after I/R., (Copyright © 2019. Published by Elsevier Ltd.)

Despite progress in use of decellularized lung scaffolds in ex vivo lung bioengineering schemes, including use of gels and other materials derived from the scaffolds, the detailed composition and functional role of extracellular matrix (ECM) proteoglycans (PGs) and their glycosaminoglycan (GAG) chains remaining in decellularized lungs, is poorly understood. Using a commonly utilized detergent-based decellularization approach in human autopsy lungs resulted in disproportionate losses of GAGs with depletion of chondroitin sulfate/dermatan sulfate (CS/DS) > heparan sulfate (HS) > hyaluronic acid (HA). Specific changes in disaccharide composition of remaining GAGs were observed with disproportionate loss of NS and NS2S for HS groups and of 4S for CS/DS groups. No significant influence of smoking history, sex, time to autopsy, or age was observed in native vs. decellularized lungs. Notably, surface plasmon resonance demonstrated that GAGs remaining in decellularized lungs were unable to bind key matrix-associated growth factors FGF2, HGF, and TGFβ1. Growth of lung epithelial, pulmonary vascular, and stromal cells cultured on the surface of or embedded within gels derived from decellularized human lungs was differentially and combinatorially enhanced by replenishing specific GAGs and FGF2, HGF, and TGFβ1. In summary, lung decellularization results in loss and/or dysfunction of specific GAGs or side chains significantly affecting matrix-associated growth factor binding and lung cell metabolism. GAG and matrix-associated growth factor replenishment thus needs to be incorporated into schemes for investigations utilizing gels and other materials produced from decellularized human lungs. STATEMENT OF SIGNIFICANCE: Despite progress in use of decellularized lung scaffolds in ex vivo lung bioengineering schemes, including use of gels and other materials derived from the scaffolds, the detailed composition and functional role of extracellular matrix (ECM) proteoglycans (PGs) and their glycosaminoglycan (GAG) chains remaining in decellularized lungs, is poorly understood. In the current studies, we demonstrate that glycosaminoglycans (GAGs) are significantly depleted during decellularization and those that remain are dysfunctional and unable to bind matrix-associated growth factors critical for cell growth and differentiation. Systematically repleting GAGs and matrix-associated growth factors to gels derived from decellularized human lung significantly and differentially affects cell growth. These studies highlight the importance of considering GAGs in decellularized lungs and their derivatives., (Copyright © 2019. Published by Elsevier Ltd.)

Aims: To study the effect of Adipose-derived stem cells (ADSCs) on fibrosis of hypertrophic scar-derived fibroblasts (HSFs) and its concrete mechanism., Background: ADSCs have been reported to reduce collagen production and fibroblast proliferation in co-culture experiments. Conditioned medium from adipose-derived stem cells (ADSCs-CM) has successfully inhibited fibrosis by decreasing the expression of collagen type І (Col1) and α-smooth muscle actin (α-SMA) in rabbit ear scar models. Hepatocyte growth factor (HGF), the primary growth factor in ADSCs-CM, has been shown to reverse fibrosis in various fibrotic diseases., Objective: To test the hypothesis that ADSCs inhibit fibrosis of HSFs through the secretion of HGF., Methods: HSFs were treated with DMEM containing 0%, 10%, 50% and 100% concentration of ADSCs-CM. The effect of ADSCs-CM on the viability was determined by cell viability assay, and the collagen production in HSFs was examined by Sirius red staining. Expression and secretion of fibrosis and degradation proteins were detected separately. After measuring the concentration of HGF in ADSCs-CM, the same number of HSFs were treated with 50% ADSCs-CM or HGF. HGF activity in ADSCs-CM was neutralized with a goat anti-human HGF antibody., Results: The results demonstrated that ADSCs-CM dose-dependently decreased cell viability, expression of fibrosis molecules, and tissue inhibitor of metalloproteinases-1 (TIMP-1), and significantly increased matrix metalloproteinase-1 (MMP-1) expression in HSFs. Collagen production and the ratio of collagen type І and type III (Col1/Col3) were also suppressed by ADSCs-CM in a dose-dependent manner. When HSFs were cultured with either 50% ADSCs-CM or HGF (1 ng/ml), a similar trend was observed in gene expression and protein secretion. Adding an HGF antibody to both groups returned protein expression and secretion to basal levels but did not significantly affect the fibrosis factors in the control group., Conclusion: Our findings revealed that adipose-derived stem cell-secreted HGF effectively inhibits fibrosis-related factors and regulates extracellular matrix (ECM) remodeling in hypertrophic scar fibroblasts., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background and Objective: Endothelial-mesenchymal transition (EndMT) not only occurs during embryonic development, but also contributes to various diseases including cardiovascular diseases, fibrosis, and even cancer. However, the specific molecular biological mechanism and relationship of related pathways have not been fully elucidated. This study aims to explore the inhibitory effect of HGF on EndMT and the molecular mechanism of Notch signal in this process., Methods: HUVECs were treated with TGF-β1 and/or HGF for 72 hours. Expression levels of EndMT markers and the key transcriptional regulators of Notch signaling pathway were assessed by qRT-PCR and western blotting. C-Met expression was measured by qRT-PCR., Results: CD31 was downregulated and α-SMA, FSP1 were upregulated during TGF-β1-induced EndMT. HGF treatment significantly attenuates the development of TGF-β1-induced EndMT by down-regulating the signal transduction of the Notch signal pathway., Conclusion: This study proves that HGF treatment significantly attenuates the development of TGF- β1-induced EndMT by inhibiting the Notch signaling, which may provide new theoretical basis for the treatment of vascular diseases and numerous fibrotic diseases caused by EndMT., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The Met/hepatocyte growth factor (HGF) receptor tyrosine kinase (RTK) is deregulated in many cancers and is a recognized target for cancer therapies. Following HGF stimulation, the signaling output of Met is tightly controlled by receptor internalization and sorting for degradation or recycling. Here, we uncover a role for autophagy in selective degradation of Met and regulation of Met-dependent cell migration and invasion. Met engagement with the autophagic pathway is dependent on complex formation with the mammalian ATG8 family member MAP1LC3C. LC3C deletion abrogates Met entry into the autophagy-dependent degradative pathway, allowing identification of LC3C domains required for rescue. Cancer cells with low LC3C levels show enhanced Met stability, signaling, and cell invasion. These findings provide mechanistic insight into RTK recruitment to autophagosomes and establish distinct roles for ATG8 proteins in this process, supporting that differential expression of ATG8 proteins can shape the functional consequences of autophagy in cancer development and progression., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Programming cells to sense multiple inputs and activate cellular signal transduction cascades is of great interest. Although this goal has been achieved through the engineering of genetic circuits using synthetic biology tools, a nongenetic and generic approach remains highly demanded. Herein, we present an aptamer-controlled logic receptor assembly for modulating cellular signal transduction. Aptamers were engineered as "robotic arms" to capture target receptors (c-Met and CD71) and a DNA logic assembly functioned as a computer processor to handle multiple inputs. As a result, the DNA assembly brings c-Met and CD71 into close proximity, thus interfering with the ligand-receptor interactions of c-Met and inhibiting its functions. Using this principle, a set of logic gates was created that respond to DNA strands or light irradiation, modulating the c-Met/HGF signal pathways. This simple modular design provides a robust chemical tool for modulating cellular signal transduction., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Mesenchymal stem cells (MSCs) have been shown to alleviate acute lung injury (ALI) via paracrine hepatocyte growth factor (HGF) and to induce the differentiation of dendritic cells (DCs) into tolerogenic dendritic cells (DCregs) and participate in the immune response. However, whether MSCs induce the production of DCregs by secreting HGF to alleviate early ALI remains unclear. We observed that the protective effect of mouse bone marrow-derived MSCs against lipopolysaccharide (LPS)-induced ALI was achieved by inducing mature DCs (mDCs) to differentiate into DCregs, and its mechanism is related to the activation of the HGF/Akt pathway., Methods: MSCs or MSCs with overexpression or knockdown of HGF were cocultured with DCs derived from mouse bone marrow using a Transwell system for 3 days. Moreover, we used MSCs or MSCs with overexpression or knockdown of HGF to treat LPS-induced ALI mice for 24 h. Flow cytometry was performed to measure the phagocytosis, accumulation, and maturation of DCs, as well as proliferation of T cells. Lung injury was estimated by lung wet weight to body weight ratio (LWW/BW) and histopathological analysis. Furthermore, we used the Akt inhibitor MK-2206 in a coculture system to elucidate the role of the HGF/Akt pathway in regulating the differentiation of DCs into regulatory DCs and relieving lung injury in early ALI mice., Results: Immature DCs (imDCs) were induced to mature after 24 h of LPS (50 ng/ml) stimulation. MSCs or HGF induced the differentiation of mDCs into regulatory DCs characterized by low expression of MHCII, CD86, and CD40 molecules, strong phagocytic function, and the ability to inhibit T cell proliferation. The effect of MSCs on DCregs was enhanced with the increase in HGF secretion and was weakened with the decrease in HGF secretion. DCregs induced by recombinant HGF were attenuated by the Akt inhibitor MK-2206. Lung DC aggregation and mDC ratio increased in LPS-induced ALI mice, while treatment with MSCs decreased lung DC aggregation and maturation and alleviated lung pathological injury. High expression of the HGF gene enhanced the above effect of MSCs, while decreased expression of HGF weakened the above effect of MSCs., Conclusions: MSCs alleviate early ALI via paracrine HGF by inducing mDCs to differentiate into regulatory DCs. Furthermore, the mechanism of HGF-induced differentiation of mDCs into DCregs is related to the activation of the Akt pathway.

Sorafenib is an oral multikinase inhibitor that has become an established therapeutic approach in advanced hepatocellular carcinoma (HCC). However, the benefit of sorafenib in clinical therapy is often affected by drug resistance. Therefore, it is important to explore the mechanisms underlying sorafenib resistance and to develop individualized therapeutic strategies for coping with this problem. In this study, we found that addition of HGF to sorafenib-treated HCC cells activated MET and re-stimulated the downstream AKT and ERK1/2 pathways. Thereby, restored sorafenib-treated HCC cells proliferation, migration and invasion ability, and rescued cells from apoptosis. In addition, we found that HGF treatment of HCC cells induced early growth response protein (EGR1) expression, which is involved in sorafenib resistance. Importantly, the HGF rescued effect in sorafenib-treated HCC cells could be abrogated by inhibiting MET activation with PHA-665752 or by downregulating EGR1 expression with small interfering RNA (siRNA). Therefore, inhibition of the HGF/MET pathway may improve response to sorafenib in HCC, and combination therapy should be further investigated.

Amyotrophic lateral sclerosis (ALS) is an adult onset neurodegenerative disorder characterized by the death of upper and lower motor neurons. About 10% of all ALS cases are familial, and we have identified SOD1 and FUS mutations as the most common causes in a consecutive series of 111 familial ALS pedigrees in Japan (Nishiyama A, 2017). From studies of the TDP43, FUS, and C9orf72 genes, perturbations of RNA processing can be highly adverse in motor neurons. Hepatocyte growth factor (HGF) is one of the most potent survival-promoting factors for motor neurons. We administered human recombinant HGF (hrHGF) by continuous intrathecal delivery to transgenic rats at onset of paralysis for 4 weeks. Intrathecal administration of hrHGF attenuated motor neuron degeneration and prolonged the duration of the disease by 63% (Ishigaki A, 2007). To translate this strategy to human treatment, we induced a contusive cervical spinal cord injury in the common marmoset, a primate, and then administered hrHGF intrathecally. We conducted a first-in-human phase I trial of intrathecal hrHGF in 15 Japanese patients with ALS (Warita H, 2019). Based on the results, we are conducting a phase II trial of intrathecal hrHGF for patients with ALS.

Treatment of muscle-invasive bladder cancer remains a major clinical challenge. Aberrant HGF/c-MET upregulation and activation is frequently observed in bladder cancer correlating with cancer progression and invasion. However, the mechanisms underlying HGF/c-MET-mediated invasion in bladder cancer remains unknown. As part of a negative feedback loop SMAD7 binds to SMURF2 targeting the TGFβ receptor for degradation. Under these conditions, SMAD7 acts as a SMURF2 agonist by disrupting the intramolecular interactions within SMURF2. We demonstrate that HGF stimulates TGFβ signalling through c-SRC-mediated phosphorylation of SMURF2 resulting in loss of SMAD7 binding and enhanced SMURF2 C2-HECT interaction, inhibiting SMURF2 and enhancing TGFβ receptor stabilisation. This upregulation of the TGFβ pathway by HGF leads to TGFβ-mediated EMT and invasion. In vivo we show that TGFβ receptor inhibition prevents bladder cancer invasion. Furthermore, we make a rationale for the use of combinatorial TGFβ and MEK inhibitors for treatment of high-grade non-muscle-invasive bladder cancers.

Objective: Hepatocyte growth factor (HGF) and its receptor MET are expressed in the salivary glands during developmental stages and tumor formation; however, the function of HGF in injured salivary gland tissues remains unclear. The present study investigated the role of HGF in protecting the salivary glands against radiation-induced injury using an organotypic culture method., Materials and Methods: Acinar-like organoids were formed by means of a three-dimensional (3D) human parotid tissue-derived spheroids (hPTS) culture method. Radioprotective effects of HGF on irradiated hPTS and signaling pathways on radioprotection were investigated., Results: We detected MET expression in hPTS grown in a 3D culture. Treatment of irradiated hPTS with recombinant human HGF (rhHGF) restored salivary marker expression and secretory function of hPTS. Changes in the phosphorylation levels of apoptosis-related proteins through HGF-MET axis inhibited radiation-induced apoptosis. Treatment with PHA665752, a MET inhibitor, blocked MET-PI3K-AKT pathway, increased apoptosis, and suppressed the radioprotective effect of rhHGF against IR-induced damage of hPTS., Conclusions: These results suggest that HGF is a key effector of radioprotection and that HGF-MET-PI3K-AKT axis is involved in protecting the salivary glands from radiation-induced apoptosis., (Copyright © 2019 The Authors. Published by Elsevier B.V. All rights reserved.)

Immune checkpoint inhibitors targeting programmed cell death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1) have improved the survival of patients with non-small cell lung cancer (NSCLC). Still, many patients do not respond to these inhibitors. PD-L1 (CD274) expression, one of the factors that influences the efficacy of immune checkpoint inhibitors, is dynamic. Here, we studied the regulation of PD-L1 expression in NSCLC without targetable genetic alterations in EGFR, ALK, BRAF, ROS1, MET, ERBB2 and RET. Analysis of RNA sequencing data from these NSCLCs revealed that inferred IFNγ, EGFR and MAPK signaling correlated with CD274 gene expression in lung adenocarcinoma. In a representative lung adenocarcinoma cell line panel, stimulation with EGF or IFNγ increased CD274 mRNA and PD-L1 protein and membrane levels, which were further enhanced by combining EGF and IFNγ. Similarly, tumor cell PD-L1 membrane levels increased after coculture with activated peripheral blood mononuclear cells. Inhibition of the MAPK pathway, using EGFR inhibitors cetuximab and erlotinib or the MEK 1 and 2 inhibitor selumetinib, prevented EGF- and IFNγ-induced CD274 mRNA and PD-L1 protein and membrane upregulation, but had no effect on IFNγ-induced MHC-I upregulation. Interestingly, although IFNγ increases transcriptional activity of CD274, MAPK signaling also increased stabilization of CD274 mRNA. In conclusion, MAPK pathway activity plays a key role in EGF- and IFNγ-induced PD-L1 expression in lung adenocarcinoma without targetable genetic alterations and may present a target to improve the efficacy of immunotherapy. © 2019 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland., (© 2019 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.)

Heterotopic ossification is common in tendon healing after trauma, but the detailed mechanisms remain unknown. Tendon-derived stem cells (TDSCs) are a type of progenitor cell found in the tendon niche, and their incorrect differentiation after trauma may lead to tendon calcification. The expression of hepatocyte growth factor (HGF) presents drastic fluctuations in serum/tissue after trauma and was found to activate quiescent stellate cells and contribute to wound healing; however, its potential role in TDSCs remains elusive. In this study, TDSCs isolated from rats were cultured in media containing HGF with or without a signaling inhibitor, and the proliferation, migration, and differentiation ability of TDSCs were measured to determine the role and mechanism of HGF in TDSCs. We showed that HGF promotes TDSC proliferation and migration but inhibits TDSC osteogenic differentiation ability. HGF activated-HGF/c-Met, mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinases 1 and 2 (ERK1/2), and phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling, which was positively correlated with TDSCs proliferation and migration but negatively related to TDSC osteogenic differentiation ability. The phosphorylation of Smad1/5/8 was also negatively related to HGF/c-Met, MAPK/ERK1/2, and PI3K/AKT signaling, which demonstrated that the inhibition of osteogenic differentiation was dependent on BMP/Smad1/5/8 signaling. Overall, we showed that HGF could promote TDSCs proliferation and migration and inhibit osteogenic differentiation in vitro, suggesting a potential role for HGF as a cytokine treatment of tendon trauma., (© 2019 Wiley Periodicals, Inc.)

Transforming growth factor-β (TGF-β)-induced fibroblast-to-myofibroblast differentiation contributes to remodeling in chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis, but whether this impacts the ability of fibroblasts to support lung epithelial repair remains little explored. We pretreated human lung fibroblasts [primary (phFB) or MRC5 cells] with recombinant human TGF-β to induce myofibroblast differentiation, then cocultured them with adult mouse lung epithelial cell adhesion molecule-positive cells (EpCAM + ) to investigate their capacity to support epithelial organoid formation in vitro. While control phFB and MRC5 lung fibroblasts supported organoid formation of mouse EpCAM + cells, TGF-β pretreatment of both phFB and MRC5 impaired organoid-supporting ability. We performed RNA sequencing of TGF-β-treated phFB, which revealed altered expression of key Wnt signaling pathway components and Wnt/β-catenin target genes, and modulated expression of secreted factors involved in mesenchymal-epithelial signaling. TGF-β profoundly skewed the transcriptional program induced by the Wnt/β-catenin activator CHIR99021. Supplementing organoid culture media recombinant hepatocyte growth factor or fibroblast growth factor 7 promoted organoid formation when using TGF-β pretreated fibroblasts. In conclusion, TGF-β-induced myofibroblast differentiation results in Wnt/β-catenin pathway skewing and impairs fibroblast ability to support epithelial repair likely through multiple mechanisms, including modulation of secreted growth factors.

Epithelial cells form a globular organ-like multi-cellular structure called cyst when cultured in extracellular matrix. The cyst generates extension followed by cell chains and tubules in response to hepatocyte growth factor (HGF). The Rho family small GTPases play essential roles for tubulogenesis. FilGAP, a Rac specific Rho GTPase-activating protein, is highly expressed in kidney. In this study, we examined the role of FilGAP in the tubulogenesis of Madin-Darby Canine Kidney (MDCK) epithelial cells. HGF induces basolateral extensions from cysts. Depletion of FilGAP by siRNA increased the number of extensions in response to HGF, whereas forced expression of FilGAP decreased the number of the extensions. FilGAP is phosphorylated and activated downstream of Rho-ROCK-signaling. Overexpression of phospho-mimic FilGAP (ST/D) mutant blocked formation of the membrane extensions induced by HGF in the presence of ROCK inhibitor, Y-27632. On the other hand, treatment of the tubules with Y27632 induced scattering of the cells, but FilGAP (ST/D) blocked cell scattering and promoted lumen formation. Taken together, our study suggests that FilGAP may suppress formation of extensions whereas stabilize tubule formation downstream of Rho-ROCK-signaling., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Collective cellular migration plays a central role in development, regeneration, and metastasis. In these processes, mechanical interactions between cells are fundamental but measurement of these interactions is often hampered by technical limitations. To overcome some of these limitations, here we describe a system that integrates microfluidics with traction microscopy (TM). Using this system we can measure simultaneously, and in real time, migration speeds, tractions, and intercellular tension throughout an island of confluent Madin-Darby canine kidney (MDCK) cells. The cell island is exposed to hepatocyte growth factor (HGF) at a controlled gradient of concentrations; HGF is known to elicit epithelial-to-mesenchymal transition (EMT) and cell scattering. As expected, the rate of expansion of the cell island was dependent on the concentration of HGF. Higher concentrations of HGF reduced intercellular tensions, as expected during EMT. A novel finding, however, is that the effects of HGF concentration and its gradient were seen within an island. This integrated experimental system thus provides an integrated tool to better understand cellular forces during collective cellular migration under chemical gradients.

Hepatocyte growth factor (HGF) is an endogenously expressed bioactive substance that has a strong anti-apoptotic effect. In this study, we biochemically and histologically characterized the effects of rh-HGF on in vitro human hepatocyte injury and mouse acute liver failure (ALF) models, both of which were induced by antibody-mediated Fas signaling. rh-HGF inhibited intracellular caspase-3/7 activation and cytokeratin 18 (CK-18) fragment release in both models. Histologically, rh-HGF dramatically suppressed parenchymal damage and intrahepatic hemorrhage. Among the laboratory parameters, prothrombin time (PT) was strongly preserved by rh-HGF, and PT was well correlated with the degree of intrahepatic hemorrhage. These results showed that the anti-apoptotic effect of rh-HGF on hepatocytes coincided strikingly with the suppression of intrahepatic hemorrhage. PT was considered to be the best parameter that correlated with the intrahepatic hemorrhages associated with hepatocellular damage. The action of rh-HGF might derive not only from its anti-apoptosis effects on liver parenchymal cells but also from its stabilization of structural and vasculature integrity., Competing Interests: Akio Ido has received honoraria from Bristol-Meyers Squibb Co., Ltd., Gilead Sciences Co., Ltd., Abbvie Inc. Those funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results. Also Akio Ido has received honoraria from Eisai Co., Ltd., and research funding from Eisai Co., Ltd.

Background & Objectives: Hepatocyte growth factor (HGF) produced by endothelial cells, fibroblasts, fat cells and other interstitial cells, can promote angiogenesis, repair damaged tissues and resist fibrosis. Mesenchymal stem cells (MSCs) are located in bone marrow and secrete a variety of cytokines and are often used in the repair and regeneration of damaged tissues. This study was aimed to investigate the influence of HGF-transfected bone marrow-derived MSCs towards renal fibrosis in rats., Methods: The HGF gene-carrying adenoviral vector (Ad-HGF) was transfected into MSCs, and the Ad-HGF-modified MSCs were transplanted into rats with unilateral ureteral obstruction (UUO). The localization of renal transplanted cells in the frozen section was observed with fluorescence microscope. The Masson's trichrome staining was performed to observe the renal collagen deposition, and the immunohistochemistry was performed to detect the expressions of α-smooth muscle actin (α-SMA) and HGF in renal tissues. Reverse transcription (RT)-PCR was used to detect the mRNA expressions of α-SMA, HGF and fibronectin (FN)., Results: Ad-HGF-modified MSCs could highly express HGF in vitro. On the post-transplantation 3 rd , 7 th and 14 th day, the 4',6-diamidino-2-phenylindole (DAP)-labelled transplanted cells were seen inside renal tissues. Compared with UUO group, the renal collagen deposition in transplantation group was significantly reduced, and the expressions of α-SMA mRNA and protein were significantly decreased, while the expressions of HGF mRNA and protein were significantly increased, and the expression of FN mRNA was significantly decreased (P<0.001)., Interpretation & Conclusions: Trans-renal artery injection of HGF-modified MSCs can effectively reduce the renal interstitial fibrosis in UUO rat model., Competing Interests: None

Clinical treatment options for human cholangiocarcinoma (CC) are limited. c-MET, a high-affinity receptor for hepatocyte growth factor (HGF), is deregulated in many cancers. Its role in cholangiocarcinogenesis remains unclear. In current study, 23 corresponding tumor- and non-tumor tissues, taken from patients with intrahepatic (iCC) and perihilar cholangiocarcinoma (pCC), who underwent liver resection, were analyzed. The relationship of clinicopathological features and c-MET, as well as c-jun N-terminal kinase (JNK) was evaluated. The anti-tumor effects of Tivantinib, a small-molecule inhibitor with potent activity against the c-MET kinase, was investigated in three human CC cell lines, namely HUCC-T1, TFK-1, and EGI-1. In comparison with the results obtained in non-tumor tissue samples, c-MET was overexpressed in 91.3 % of tumor tissues (p < 0.01). The JNK expression was higher in tumor tissue compared with the corresponding non-tumor tissue sample in 17.4% patients (p < 0.01). The inhibition of aberrant c-MET expression in human CC cell lines was achieved by blocking the phosphorylation of c-MET with Tivantinib. Notable losses in cell viability and colony-forming capability were detected (p < 0.01). Synergistic activation of the JNK/c-jun pathway was demonstrated after Tivantinib treatment. Knockdown of the JNK by siRNA or competitive binding of c-MET receptor by stimulation with HGF-antagonized anti-tumor effects of Tivantinib was observed. Our data suggest that inhibition of c-MET could be a possible alternative approach for the treatment of human CC, for which Tivantinib may an effective inhibitor. The synergistic activation of the JNK/c-jun pathway contributed to the elevated apoptosis in CC cells via treatment with Tivantinib.

With increasing demand for treatment of glottal insufficiency, several injection materials have been examined. However, biological resorption, degradation of injected materials, and the subsequent need to perform multiple injections still remain major clinical problems. In this study, we fabricated two different growth factor (GF) [single basic fibroblast growth factor (bFGF), single hepatocyte growth factor (HGF), or dual bFGF/HGF]-immobilized polycaprolactone (PCL)/Pluronic F127 microspheres. These materials were investigated for their potential use as bioactive injection laryngoplasty agents. HGF was found to be continuously released over 20 days and the bFGF was found to be continuously released over 25 days, as demonstrated by ELISA assay. Human vocal fold fibroblasts (hVFFs) showed significantly higher proliferative ability on dual GF-immobilized microspheres. GF-immobilized microspheres (bFGF, HGF, and dual GF) were injected into paralyzed vocal folds of New Zealand white rabbits. Through endoscopic observation and H&E staining, we identified that the microspheres remained localized at the injection site, resulting in constant volume augmentation of the paralyzed vocal fold without significant loss of the initial volume after 4 weeks. The expression of genes related to the extracellular matrix (ECM) in the vocal fold was upregulated by dual GF-immobilized microspheres. Furthermore, dual GF-immobilized microspheres inhibited muscle degeneration and upregulation of myogenic-related genes. In conclusion, dual GF-immobilized microspheres passively augmented the volume of the paralyzed vocal fold while actively inducing ECM synthesis at the injected vocal fold and preserving muscle tissue. Dual GF-immobilized microspheres could be a new and promising injection material for paralyzed vocal folds. STATEMENT OF SIGNIFICANCE: Limitation of prolonged augmentation of vocal fold and degeneration of vocal fold tissue still remain as major clinical problems in the treatment of vocal fold paralysis. Herein, we fabricated the polycaprolactone (PCL)/Pluronic F127 microspheres to augment volume of paralyzed vocal folds. On top of that, we additionally immobilized the growth factors (bFGF, HGF, or dual bFGF/HGF) on the surface of these microspheres. We highlight the efficacy of the dual GF-immobilized microspheres which augmented the volume of the paralyzed vocal fold passively, induced ECM synthesis actively at the injected vocal fold and preserved laryngeal muscle tissue. Our results suggest that the dual GF-immobilized microsphere could be a new promising injection material for injection laryngoplasty to treat paralyzed vocal fold., (Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Objective: To investigate role of β-catenin and lncRNA MALAT1/miR-217 axis to converge into the regulation of ZEB-1 in hepatocyte growth factor (HGF)-induced hepatocytes differentiated from bone marrow mesenchymal stem cells (BM-MSCs)., Methods: BM-MSCs were isolated and HGF was used to induce the differentiation of BM-MSCs into hepatocytes. HSC-T6 cells, BRL-3 A cells and differentiated BM-MSCs were treated by lipopolysaccharide(LPS). shRNAs were used to silence β-catenin and recombinant plasmids were used to over-express ZEB1. Measurement of cell viability was conducted using MTT assay and Hoechst 33342 staining. RNA immunoprecipitation (RIP) assay was used to determine binding of miR-217-3p and MALAT1., Results: BM-MSCs successfully differentiated into hepatocytes by HGF treatment. Expression of β-catenin, ZEB-1 and TERT was up-regulated to a higher level in hepatocytes differentiated from BM-MSCs than HSC-T6 cells and BRL-3 A cells after LPS stimulation. When β-catenin was knocked down in all cell lines, expression of β-catenin, ZEB-1 and TERT was significantly decreased as well as telomerase activity. While when ZEB1 was over-expressed, expression of TERT and telomerase activity was all significantly up-regulated. In hepatocytes differentiated from BM-MSCs, miR-217 was down-regulated and lncRNA MALAT1 was up-regulated. RIP analysis showed MALAT1 was physically associated with miR-217 and might function in the regulation of ZEB-1, further enhancing the expression of TERT so as to augment telomerase activity., Conclusion: We successfully used HGF to mediate differentiation of BM-MSCs into hepatocytes, and found that β-catenin-coordinated MALAT1/miR-217 axis could up-regulate expression of ZEB-1 and further enhanced the telomerase activity through regulation of TERT in BM-MSCs differentiating into hepatocytes., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Objectives/hypothesis: To elucidate the aging physiology of the vocal folds, we examined the characters of aged vocal fold fibroblasts (VFFs) in various conditions., Study Design: In vitro study., Methods: VFFs from young (12-week-old) and aged (19-month-old) Sprague-Dawley rats were compared. Proliferative capacity, ratio of myofibroblast to fibroblast, myofibroblast function, and extracellular matrix production were examined in the following conditions: naïve, basic fibroblast growth factor (bFGF) supplemented, and hepatocyte growth factor (HGF) supplemented., Results: Aged VFFs demonstrated reduced proliferation by cell counting, though the ratio of Ki-67-positive cells showed no difference. Aged VFFs exhibited an increased expression of α-smooth muscle actin (α-SMA); however, they demonstrated no enhanced contractile ability in a gel contraction assay. Type I collagen protein was increased age dependently, accompanied with decreased Mmp1 and unchanged Col1a1 transcription. Type I collagen protein and α-SMA represented quite similar reduction patterns to bFGF or HGF administration., Conclusions: The following possible characteristics of aged VFFs were implied: long duration of mitosis, increased myofibroblast population size with certain dysfunctions, reduced type I collagen turnover, and correlation between α-SMA expression and type I collagen metabolism. Further investigations of these features will help to clarify presbyphonia's pathology and establish treatment strategies., Level of Evidence: NA Laryngoscope, 129:E94-E101, 2019., (© 2018 The American Laryngological, Rhinological and Otological Society, Inc.)

Inadequate migration and invasion of the trophoblast cells during embryo implantation is one of the reasons for pregnancy-related complications such as intrauterine growth restriction and preeclampsia. In the present study, relevance of WNT ligands and integrins associated with hepatocyte growth factor (HGF)-mediated migration of HTR-8/SVneo trophoblastic cells has been investigated. Treatment of HTR-8/SVneo cells with HGF led to a dose-dependent increase in their migration. RT-PCR studies revealed a significant increase in the transcripts of WNT4, WNT11, ITGA2, and ITGAV, which was further confirmed at protein level by Western blotting. HGF treatment also led to increased expression of integrin α2β1 and αVβ5 in HTR-8/SVneo cells. Silencing of WNT4, WNT11, ITGA2, and ITGAV by siRNA led to a significant decrease in HGF-mediated migration of cells. Treatment of cells with HGF led to activation of mitogen-activated protein kinases (MAPK) and protein kinase A (PKA) signaling pathways. Inhibition of MAPK/PKA, by selective inhibitors, led to decrease in the expression of above WNT ligands and integrins. Silencing of WNT4/WNT11 led to concomitant decrease in the expression of ITGA2 and ITGAV and vice versa. HGF treatment also led to significant increase in β-catenin expression, a downstream target of both WNT ligands and integrins. Silencing of β-catenin led to decrease in HGF-mediated migration. β-catenin expression was also down-regulated in WNT4/WNT11/ITGA2/ITGAV silenced cells suggesting a possible cross-communication of WNT ligands and integrins via β-catenin. These studies have established the significance of WNT4/WNT11 as well as ITGA2/ITGAV during HGF-mediated migration of HTR-8/SVneo trophoblastic cells.