5,7,3',4'-Tetramethoxyflavone suppresses TGF-ß1-induced activation of murine fibroblasts in vitro and ameliorates bleomycin-induced pulmonary fibrosis in mice.

OBJECTIVE: This study aimed to investigate the use of 5,7,3',4'-tetramethoxyflavone (TMF) to treat pulmonary fibrosis (PF), a chronic and fatal lung disease. In vitro and in vivo models were used to examine the impact of TMF on PF. METHODS: NIH-3T3 (Mouse Embryonic Fibroblast) were exposed to transforming growth factor­ß1 (TGF-ß1) and treated with or without TMF. Cell growth was assessed using the MTT method, and cell migration was evaluated with the scratch wound assay. Protein and messenger ribonucleic acid (mRNA) levels of extracellular matrix (ECM) genes were analyzed by western blotting and quantitative reverse transcription-polymerase chain reaction (RT-PCR), respectively. Downstream molecules affected by TGF-ß1 were examined by western blotting. In vivo, mice with bleomycin-induced PF were treated with TMF, and lung tissues were analyzed with staining techniques. RESULTS: The in vitro results showed that TMF had no significant impact on cell growth or migration. However, it effectively inhibited myofibroblast activation and ECM production induced by TGF-ß1 in NIH-3T3 cells. This inhibition was achieved by suppressing various signaling pathways, including Smad, mitogen-activated protein kinase (MAPK), phosphoinositide 3-kinase/AKT (PI3K/AKT), and WNT/ß-catenin. The in vivo experiments demonstrated the therapeutic potential of TMF in reducing PF induced by bleomycin in mice, and there was no significant liver or kidney toxicity observed. CONCLUSION: These findings suggest that TMF has the potential to effectively inhibit myofibroblast activation and could be a promising treatment for PF. TMF achieves this inhibitory effect by targeting TGF-ß1/Smad and non-Smad pathways.

Mesenteric lymph nodes: a critical site for the up-regulatory effect of hUC-MSCs on Treg cells by producing TGF-ß1 in colitis treatment.

BACKGROUND: Mesenchymal stem cells (MSCs) demonstrate a wide range of therapeutic capabilities in the treatment of inflammatory bowel disease (IBD). The intraperitoneal injection of MSCs has exhibited superior therapeutic efficacy on IBD than intravenous injection. Nevertheless, the precise in vivo distribution of MSCs and their biological consequences following intraperitoneal injection remain inadequately understood. Additional studies are required to explore the correlation between MSCs distribution and their biological effects. METHODS: First, the distribution of human umbilical cord MSCs (hUC-MSCs) and the numbers of Treg and Th17 cells in mesenteric lymph nodes (MLNs) were analyzed after intraperitoneal injection of hUC-MSCs. Subsequently, the investigation focused on the levels of transforming growth factor beta1 (TGF-ß1), a key cytokine to the biology of both Treg and Th17 cells, in tissues of mice with colitis, particularly in MLNs. The study also delved into the impact of hUC-MSCs therapy on Treg cell counts in MLNs, as well as the consequence of TGFB1 knockdown hUC-MSCs on the differentiation of Treg cells and the treatment of IBD. RESULTS: The therapeutic effectiveness of intraperitoneally administered hUC-MSCs in the treatment of colitis was found to be significant, which was closely related to their quick migration to MLNs and secretion of TGF-ß1. The abundance of hUC-MSCs in MLNs of colitis mice is much higher than that in other organs even the inflamed sites of colon. Intraperitoneal injection of hUC-MSCs led to a significant increase in the number of Treg cells and a decrease in Th17 cells especially in MLNs. Furthermore, the concentration of TGF-ß1, the key cytokine for Treg differentiation, were also found to be significantly elevated in MLNs after hUC-MSCs treatment. Knockdown of TGFB1 in hUC-MSCs resulted in a noticeable reduction of Treg cells in MLNs and the eventually failure of hUC-MSCs therapy in colitis. CONCLUSIONS: MLNs may be a critical site for the regulatory effect of hUC-MSCs on Treg/Th17 cells and the therapeutic effect on colitis. TGF-ß1 derived from hUC-MSCs promotes local Treg differentiation in MLNs. This study will provide new ideas for the development of MSC-based therapeutic strategies in IBD patients.

Low frequency of repetitive trans-spinal magnetic stimulation promotes functional recovery after spinal cord injury in mice through inhibiting TGF-ß1/Smad2/3 signaling pathway.

Spinal cord injury (SCI) remains a worldwide challenge due to limited treatment strategies. Repetitive trans-spinal magnetic stimulation (rTSMS) is among the most cutting-edge treatments for SCI. However, the mechanism underlying rTSMS on functional recovery is still unclear. In this study, 8-week-old C57BL/6J female mice were used to design SCI models followed by treatment with monotherapy (1 Hz rTSMS or LY364947) or combination therapy (rTSMS + LY364947). Our results showed obvious functional recovery after monotherapies compared to untreated mice. Immunofluorescence results demonstrated that rTSMS and LY364947 modulate the lesion scar by decreasing fibrosis and GFAP and possess the effect on neural protection. In addition, rTSMS suppressed inflammation and the activation of TGFß1/Smad2/3 signaling pathway, as evidenced by markedly reduced TGF-ßRⅠ, Smad2/3, and p-Smad2/3 compared with untreated mice. Overall, it was confirmed that 1 Hz rTSMS promotes SCI recovery by suppressing the TGFß1/Smad2/3 signaling, revealing a novel pathological mechanism of 1 Hz rTSMS intervention, and may provide potential targets for clinical treatment.

Caloric and time restriction diets improve acute kidney injury in experimental menopausal rats: role of silent information regulator 2 homolog 1 and transforming growth factor beta 1.

BACKGROUND: Estrogen has a protective impact on acute kidney injury (AKI); moreover, reducing the daily intake of calories impedes developing diseases. The present study aimed to determine the effects of calorie restriction (CR) and time restriction (TR) diets on the expression of silent information regulator 2 homolog 1 (SIRT1), transforming growth factor beta 1 (TGF-ß1), and other indicators in the presence and absence of ovaries in AKI female rats. METHODS: The female rats were divided into two groups, ovariectomized (OVX) and sham, and were placed on CR and TR diets for eight weeks; afterward, AKI was induced by injecting glycerol, and kidney injury indicators and biochemical parameters were measured before and after AKI. RESULTS: After AKI, the levels of urine albumin excretion rate, urea, and creatinine in serum, and TGF-ß1 increased, while creatinine clearance and SIRT1 decreased in kidney tissue. CR improved kidney indicators and caused a reduction in TGF-ß1 and an increase in SIRT1 in ovary-intact rats. Moreover, CR prevented total antioxidant capacity (TAC) decrease and malondialdehyde (MDA) increase resulting from AKI. Before AKI, an increase in body weight, fasting blood sugar (FBS), low-density lipoprotein (LDL), triglyceride (TG), and total cholesterol (TC), and a decrease in high-density lipoprotein (HDL) were observed in OVX rats compared to sham rats, but CR prevented these changes. The effects of TR were similar to those of CR in all indicators except for TGF-ß1, SIRT1, urea, creatinine, and albumin. CONCLUSION: The present study indicated that CR is more effective than TR in preventing AKI, probably by increasing SIRT1 and decreasing TGF-ß1 in ovary-intact animals.

PAR2 activation on human tubular epithelial cells engages converging signaling pathways to induce an inflammatory and fibrotic milieu.

Key features of chronic kidney disease (CKD) include tubulointerstitial inflammation and fibrosis. Protease activated receptor-2 (PAR2), a G-protein coupled receptor (GPCR) expressed by the kidney proximal tubular cells, induces potent proinflammatory responses in these cells. The hypothesis tested here was that PAR2 signalling can contribute to both inflammation and fibrosis in the kidney by transactivating known disease associated pathways. Using a primary cell culture model of human kidney tubular epithelial cells (HTEC), PAR2 activation induced a concentration dependent, PAR2 antagonist sensitive, secretion of TNF, CSF2, MMP-9, PAI-1 and CTGF. Transcription factors activated by the PAR2 agonist 2F, including NFκB, AP1 and Smad2, were critical for production of these cytokines. A TGF-ß receptor-1 (TGF-ßRI) kinase inhibitor, SB431542, and an EGFR kinase inhibitor, AG1478, ameliorated 2F induced secretion of TNF, CSF2, MMP-9, and PAI-1. Whilst an EGFR blocking antibody, cetuximab, blocked PAR2 induced EGFR and ERK phosphorylation, a TGF-ßRII blocking antibody failed to influence PAR2 induced secretion of PAI-1. Notably simultaneous activation of TGF-ßRII (TGF-ß1) and PAR2 (2F) synergistically enhanced secretion of TNF (2.2-fold), CSF2 (4.4-fold), MMP-9 (15-fold), and PAI-1 (2.5-fold). In summary PAR2 activates critical inflammatory and fibrotic signalling pathways in human kidney tubular epithelial cells. Biased antagonists of PAR2 should be explored as a potential therapy for CKD.

Kindlin-2 mediates Peyronie's disease through activation of TGF-ß/Smad signaling pathway under the presence of TGF-ß1.

BACKGROUND: Peyronie's disease (PD) causes benign plaques or induration in tunica albuginea (TA). Kindlin-2 regulates the TGF-ß1/Smad3 pathway, which accelerates kidney fibrosis. The study is aimed mainly to investigate the impact of Kindlin-2 on PD formation and its signaling pathways, notably the TGF-ß/Smad pathway in the presence of TGF-ß1. METHODS: In this mouse investigation, adenovirus TGF-ß1 was injected into TA to produce PD. The model was successfully induced 45 days later. Western Blot (WB) and immunohistochemistry (IHC) were utilized to measure Kindlin-2 in PD model tissue. WB and immunofluorescence assays were utilized to confirm the impact of TGF-ß1 on Kindlin-2 levels in vitro. The interaction among Kindlin-2, TßRI, and Smad3 was detected using immunoprecipitation (IP) experiments. We examined how TGF-ß1 affects Smad3 phosphorylation and downstream gene activation process. Finally, Kindlin-2 and the level of tissue fibrosis were examined in PD model. RESULTS: Kindlin-2 levels were elevated in the TGF-ß1-induced PD model, confirming that TGF-ß1 can increase Kindlin-2 levels in primary PD cells. Moreover, Kindlin-2 mediates Smad3-TßRI interaction, activates p-Smad3, and enhances TGF-ß1 target gene expression. In vivo investigations reveal that Kindlin-2 promotes PD development and tissue fibrosis. The regulatory effects of Kindlin-2 need the presence of TGF-ß1. Tissue fibrosis can be reduced by downregulating Kindlin-2. CONCLUSION: Kindlin-2 does not directly activate Smad3 to induce tissue fibrosis. Instead, it exerts its effect through the combined influence of TGF-ß1. Inhibiting Kindlin-2 could potentially be a treatment for PD.

Thyroid dysfunction in Hashimoto's thyroiditis: a pilot study on the putative role of miR-29a and TGFß1.

PURPOSE: Hashimoto's thyroiditis (HT) is one of the most common causes of thyroid dysfunction in iodine sufficient worldwide areas, but its molecular mechanisms are not completely understood. To this regard, this study aimed to assess serum levels of miRNA-29a (miR-29a) and transforming growth factor beta 1 (TGFß1) in HT patients with different patterns of thyroid function. METHODS: A total of 29 HT patients, with a median age of 52 years (21-68) were included. Of these, 13 had normal thyroid function (Eu-HT); 8 had non-treated hypothyroidism (Hypo-HT); 8 had hypothyroidism on replacement therapy with LT4 (subst-HT). All patients had serum miR-29a assayed through qRT-PCR and serum TGFß1 assayed by ELISA. RESULTS: Serum miR-29a levels were significantly down-regulated in patients with Hypo-HT compared to Eu-HT patients (P < 0.01) and subst-HT patients (P < 0.05). A significant negative correlation was detected between serum miR-29a levels and TSH levels (r = -0.60, P < 0.01). Serum TGFß1 levels were significantly higher in Hypo-HT than both Eu-HT (P < 0.01) and subst-HT patients (P < 0.05). A negative correlation was observed between serum miR-29a and TGFß1 (r = -0.75, P < 0.01). CONCLUSIONS: In conclusion, Hypo-HT patients had lower levels of serum miR-29a and higher levels of TGFß1 in comparison with Eu-HT patients. Worthy of note, subst-HT patients showed restored serum miR-29a levels compared with Hypo-HT group, associated with lower serum TGFß1. These novel findings may suggest a possible impact of replacement therapy with levothyroxine on serum miR-29a levels in HT.

TGF-ß1-Induced LINC01094 promotes epithelial-mesenchymal transition in hepatocellular carcinoma through the miR-122-5p/TGFBR2-SAMD2-SMAD3 Axis.

Hepatocellular carcinoma (HCC) is a common malignancy with a poor prognosis. It has been proven that long non-coding RNAs (lncRNAs) play an essential role in regulating HCC progression. However, the involvement of LINC01094 in regulating epithelial-mesenchymal transition (EMT) in HCC remains unclear. LINC01094 expression in HCC patients was retrieved from the Cancer Genome Atlas database. Overexpressing and downregulating LINC01094 were conducted to investigate its biological functions using Hep3B, SNU-387, and HuH-7 cells. Western blotting and morphological observation were performed to study the EMT in HCC cells. Transwell assay was adopted to determine the migration and invasion of HCC cells. The underlying mechanism of competitive endogenous RNAs (ceRNAs) was investigated using bioinformatics analysis, quantitative reverse-transcription polymerase chain reaction, and rescue experiments. Elevated LINC01094 expression was observed in HCC and associated with a poor prognosis. Knockdown of LINC01094 expression in SNU-387 and HuH-7 cells could inhibit migration, invasion, and EMT markers. Overexpression of LINC01094 indicated that LINC01094 promoted EMT via the TGF-ß/SMAD signaling pathway. The bioinformatics analysis revealed that miR-122-5p was a target of LINC01094. The miRWalk database analysis showed that TGFBR2, SMAD2, and SMAD3 were downstream targets of miR-122-5p. Mechanically, LINC01094 acted as a ceRNA that facilitated HCC metastasis by sponging miR-122-5p to regulate the expression of TGFBR2, SMAD2, and SMAD3. Further, TGF-ß1 could enhance the expression of LINC01094, forming a positive feedback loop. TGF-ß1-induced LINC01094 expression promotes HCC cell migration and invasion by targeting the miR-122-5p/TGFBR2-SMAD2-SMAD3 axis. LINC01094 may be a potential prognostic biomarker and therapeutic target for HCC metastasis.

The positive role of transforming growth factor-ß1 in ischemic stroke.

Ischemic stroke is one of the most disabling and fatal diseases around the world. The damaged brain tissues will undergo excessive autophagy, vascular endothelial cells injury, blood-brain barrier (BBB) impairment and neuroinflammation after ischemic stroke. However, there is no unified viewpoint on the underlying mechanism of brain damage. Transforming growth factor-ß1 (TGF-ß1), as a multi-functional cytokine, plays a crucial role in the intricate pathological processes and helps maintain the physiological homeostasis of brain tissues through various signaling pathways after ischemic stroke. In this review, we summarize the protective role of TGF-ß1 in autophagic flux, BBB, vascular remodeling, neuroinflammation and other aspects after ischemic stroke. Based on the review, we believe that TGF-ß1 could serve as a key target for treating ischemic stroke.

Mechanistic investigation of wogonin in delaying the progression of endothelial mesenchymal transition by targeting the TGF-ß1 pathway in pulmonary hypertension.

Pulmonary hypertension (PH) is characterized by pulmonary vascular remodeling, which endothelial-to-mesenchymal transition (EndMT) being its main progressive phase. Wogonin, a flavonoid extracted from the root of Scutellaria baicalensis Georgi, hinders the abnormal proliferation of cells and has been employed in the treatment of several cardiopulmonary diseases. This study was designed to investigate how wogonin affected EndMT during PH. Monocrotaline (MCT) was used to induce PH in rats. Binding capacity of TGF-ß1 receptor to wogonin detected by molecular docking and molecular dynamics. EndMT model was established in pulmonary microvascular endothelial cells (PMVECs) by transforming growth factor beta-1 (TGF-ß1). The result demonstrated that wogonin (20 mg/kg/day) attenuated right ventricular systolic pressure (RVSP), right ventricular hypertrophy and pulmonary vascular thickness in PH rats. EndMT in the pulmonary vascular was inhibited after wogonin treatment as evidenced by the restored expression of CD31 and decreased expression of α-SMA. Wogonin has strong affinity for both TGFBRI and TGFBRII, and has a better binding stability for TGFBRI. In TGF-ß1-treated PMVECs, wogonin (0.3, 1, and 3 µM) exhibited significant inhibitory effects on this transformation process via down-regulating the expression of p-Smad2 and Snail, while up-regulating the expression of p-Smad1/5. Additionally, results of Western blot and fluorescence shown that the expression of α-SMA were decrease with increasing level of CD31 in PMVECs. In conclusion, our research showed that wogonin suppressed EndMT via the TGF-ß1/Smad pathway which may lead to its alleviated effect on PH. Wogonin may be a promising drug against PH.

Longitudinal Correlations between Molecular Compositions of Stratum Corneum and Breast Milk Factors during Infancy: A Prospective Birth Cohort Study.

Breast milk contains numerous factors that are involved in the maturation of the immune system and development of the gut microbiota in infants. These factors include transforming growth factor-ß1 and 2, immunoglobin A, and lactoferrin. Breast milk factors may also affect epidermal differentiation and the stratum corneum (SC) barrier in infants, but no studies examining these associations over time during infancy have been reported. In this single-center exploratory study, we measured the molecular components of the SC using confocal Raman spectroscopy at 0, 1, 2, 6, and 12 months of age in 39 infants born at our hospital. Breast milk factor concentrations from their mothers' breast milk were determined. Correlation coefficients for the two datasets were estimated for each molecular component of the SC and breast milk factor at each age and SC depth. The results showed that breast milk factors and molecular components of the SC during infancy were partly correlated with infant age in months and SC depth, suggesting that breast milk factors influence the maturation of the SC components. These findings may improve understanding of the pathogenesis of skin diseases associated with skin barrier abnormalities.

TGF-ß1 inhibitor enhances the therapeutic effect of microwave ablation on hepatocellular carcinoma.

BACKGROUND: Microwave ablation (MWA) is a widely adopted treatment technique for hepatocellular carcinoma (HCC). However, MWA alone is of limited use and has a high recurrence rate. Transforming growth factor-ß1 (TGF-ß1) is recognized as a potential therapeutic target for HCC patients. Therefore, this study was designed to investigate whether the TGF-ß1 inhibitor could increase the efficacy of MWA therapy for HCC treatment. METHODS: In vitro, HCC cells challenged with TGF-ß1 inhibitor (SB-525334), or normal saline were then heated by microwave. Methyl tetrazolium assays were performed to detect cell survival rate and half-maximal drug inhibitory concentration (IC50). Cell viability and apoptosis were detected by cell counting kit-8 assays, flow cytometry and western blotting. In vivo, the mice injected with HepG2 cells received oral gavage of SB-525334 (20 mg/kg) or normal saline and MWA at a power of 15 W. Tumor volume was recorded. Expression of Ki67 and apoptosis-related proteins were detected by immunohistochemistry and western blotting. TUNEL assays were used to detect cell death ratio. Histopathological changes were examined by hematoxylin and eosin staining. The mechanisms associated with the function of MWA combined with TGF-ß1 inhibitor in HCC development were explored by western blotting. RESULTS: Combination of MWA and SB-525334 decreased the survival rate and promoted the apoptosis of HCC cells compared with MWA alone. SB-525334 enhanced the suppressive effect of MWA on tumor growth and amplified cell apoptosis. Mechanistically, MWA collaborated with SB-525334 inhibitor inactivated the TGF-ß1/Smad2/Smad3 pathway. CONCLUSION: TGF-ß1 inhibitor enhances the therapeutic effect of MWA on HCC.

SB-525334 ameliorates renal injury in diabetic kidney disease mouse model via suppressing inflammation.

BACKGROUND: Diabetic kidney disease (DKD; also known as diabetic nephropathy) is a typical complication of diabetes mellitus characterised by renal injury due to disturbances in glucose metabolism, in which renal tubular damage caused by chronic inflammation has been shown to be closely associated with the development of end-stage renal disease (ESRD). However, there are insufficient effective therapeutic agents to halt the progression of DKD. METHODS: In the present study, we screened differential gene expression profiles associated with DKD by mining the GEO database through differential and enrichment analyses. Furthermore, systemic in vivo and in vitro experiments were designed to explore the mechanism through which the potential therapeutic agent SB-525334 improves DKD. RESULTS: SB-525334 ameliorated DKD-induced kidney injury by regulating inflammatory cytokines (TGF-ß1, IL-6, IL-10) as well as promoting the translation of M1 (iNOS) macrophage to M2 (CD206) macrophage. In addition, SB-525334 ameliorates kidney injury caused by DKD through inhibiting inflammation through regulating the expression of key proteins in the TGF-ß1 /JNK and TGF-ß1 /Smad signaling pathways. For studies in vitro, inflammation induced by LPS in vitro was inhibited significantly after the administration of SB-525334 through down-regulating pro-inflammatory cytokines, promoting macrophage conversion from M1 to M2, and inhibiting the activation of TGF-ß1 /JNK and TGF-ß1 /Smad pathways. CONCLUSIONS: These results highlight that the target compound SB-525334 could serve as a novel potential therapeutic agent and ameliorate DKD in an inflammation-inhibiting manner.

Tanshinone IIA Liposomes Treat Doxorubicin-Induced Glomerulonephritis by Modulating the Microenvironment of Fibrotic Kidneys.

Renal fibrosis plays a key role in the pathogenesis of chronic kidney disease (CKD), in which the persistent high expression of transforming growth factor ß1 (TGF-ß1) and α-smooth muscle actin (α-SMA) contributes to the progression of CKD to renal failure. In order to improve the solubility, bioavailability, and targeting of tanshinone IIA (Tan IIA), a novel targeting material, aminoethyl anisamide-polyethylene glycol-1,2-distearoyl-sn-glycero-3-phosphate ethanolamine (AEAA-PEG-DSPE, APD) modified Tan IIA liposomes (APD-Tan IIA-L) was constructed. An animal model of glomerulonephritis induced by doxorubicin in BALB/c mice was established. APD-Tan IIA-L significantly decreased blood urea nitrogen and serum creatinine (SCr), and the consequences of renal tissue oxidative stress indicators showed that APD-Tan IIA-L downregulated malondialdehyde, upregulated superoxide dismutase, catalase, and glutathione peroxidase. Masson's trichrome staining showed that the deposition of collagen in the APD-Tan IIA-L group decreased significantly. The pro-fibrotic factors (fibronectin, collagen I, TGF-ß1, and α-SMA) and epithelial-mesenchymal transition marker (N-cadherin) were significantly inhibited by APD-Tan IIA-L. By improving the microenvironment of fibrotic kidneys, APD-Tan IIA-L attenuated TGF-ß1-induced excessive proliferation of fibroblasts and alleviated oxidative stress damage to the kidney, providing a new strategy for the clinical treatment of renal fibrosis.

Ciwujianoside E inhibits Burkitt lymphoma cell proliferation and invasion by blocking ENO1-plasminogen interaction and TGF-ß1 activation.

Burkitt's lymphoma (BL) is a rare and highly aggressive B-cell non-Hodgkin lymphoma. Although the outcomes of patients with BL have greatly improved, options for patients with relapsed and refractory BL are limited. Therefore, there is an urgent need to improve BL therapeutics and to develop novel drugs with reduced toxicity. In this study, we demonstrated that enolase 1 (ENO1) is a potential novel drug target for BL treatment. We determined that ENO1 was aberrantly upregulated in BL, which was closely related to its invasiveness and poor clinical outcomes. Furthermore, using RNA interference, we demonstrated that ENO1 depletion significantly inhibited cell proliferation and invasion both in vitro and in vivo. Mechanistically, we established that ENO1 knockdown suppressed the PI3K-AKT and epithelial-mesenchymal transition (EMT) signaling pathways by reducing plasminogen (PLG) recruitment, plasmin (PL) generation, and TGF-ß1 activation. Addition of activated TGF-ß1 protein to the culture medium of shENO1 cells reversed the inhibitory effects on cell proliferation and invasion, as well as those on the PI3K-AKT and EMT signaling pathways. Notably, our research led to the discovery of a novel ENO1-PLG interaction inhibitor, Ciwujianoside E (L-06). L-06 effectively disrupts the interaction between ENO1 and PLG, consequently reducing PL generation and suppressing TGF-ß1 activation. In both in vitro and in vivo experiments, L-06 exerted impressive antitumor effects. In summary, our study elucidated the critical role of ENO1 in BL cell proliferation and invasion and introduced a novel ENO1 inhibitor, which holds promise for improving the treatment of patients with BL in the future.

Down-regulation of TGF-ß1 in Fibro-adipogenic Progenitors Initiates Muscle Ectopic Mineralization.

In previous studies, we have demonstrated that stress response-induced high glucocorticoid levels could be the underlying cause of traumatic heterotopic ossification (HO), and we have developed a glucocorticoid-induced ectopic mineralization (EM) mouse model by systemic administration of a high dose of dexamethasone (DEX) to animals with muscle injury induced by cardiotoxin injection. In this model, dystrophic calcification (DC) developed into HO in a cell autonomous manner. However, it is not clear how DC is formed after DEX treatment. Therefore, in this study, we aimed to explore how glucocorticoids initiate muscle EM at a cellular and molecular level. We showed that DEX treatment inhibited inflammatory cell infiltration into injured muscle but inflammatory cytokine production in the muscle was significantly increased, suggesting that other non-inflammatory muscle cell types may regulate the inflammatory response and the muscle repair process. Accompanying this phenotype, transforming growth factor ß1 (TGF-ß1) expression in fibro-adipogenic progenitors (FAPs) was greatly down-regulated. Since TGF-ß1 is a strong immune suppressor and FAP's regulatory role has a large impact on muscle repair, we hypothesized that down-regulation of TGF-ß1 in FAPs after DEX treatment resulted in this hyperinflammatory state and subsequent failed muscle repair and EM formation. To test our hypothesis, we utilized a transgenic mouse model to specifically knock out Tgfb1 gene in PDGFRα positive FAPs to investigate if the transgenic mice could recapitulate the phenotype that was induced by DEX treatment. Our results showed that the transgenic mice completely phenocopied this hyperinflammatory state and spontaneously developed EM following muscle injury. On the contrary, therapeutics that enhanced TGF-ß1 signaling in FAPs inhibited the inflammatory response and attenuated muscle EM. In summary, these results indicate that FAPs-derived TGF-ß1 is a key molecule in regulating muscle inflammatory response and subsequent EM, and that glucocorticoids exert their effect via down-regulating TGF-ß1 in FAPs.

Heterotopic ossification (HO) is abnormal bone formation in soft tissue. Glucocorticoids, which have strong anti-inflammatory properties, have usually been used as HO therapeutics. However, our findings suggest that glucocorticoids can also promote HO formation. In this study, we tried to explain the underlying reason for these seemingly contradictory observations. We showed that glucocorticoids, in addition to exerting an anti-inflammatory effect on inflammatory cells, can also target another type of muscle cell to exert a pro-inflammatory effect. These cells are called fibro-adipogenic progenitors (FAPs), and we demonstrated that FAPs played a master regulatory role in the muscle inflammatory response by modulating the expression of transforming growth factor ß1 (TGF-ß1), a well-known immune suppressor. In summary, our findings highlighted the importance of FAP TGF-ß1 levels in affecting the progression and regression of muscle HO, and provided new treatment options for HO based on their ability to elevate TGF-ß1 levels in FAPs.

Effect of Calcium Silicate-Based Intracanal Medicament and Calcium Hydroxide on Growth Factor TGF-ß1 Release from Root Canal Dentine.

Effective root canal disinfection and the subsequent release of natural growth factors from dentin are crucial to the success of regenerative endodontic procedures. This study evaluated the effect of newly introduced calcium silicate-based temporary intracanal medicament Bio-C Temp and calcium hydroxide-based material UltraCal XS on the release of transforming growth factor ß1 (TGF-ß1) from root canal dentin. Twenty-two intact and fully developed human premolars from patients aged 15-18 were shaped and irrigated according to the current clinical recommendations. The teeth were then gently split in half, and the root canal dentin of paired samples was covered with Bio-C Temp or UltraCal XS. After 3 weeks of incubation, the specimens were conditioned with 17% EDTA and the collected solution was subjected to the quantification of the released TGF-ß1 by performing an ELISA. One-way analysis of variance (ANOVA), followed by Tukey's test, was selected to determine the statistically significant differences between the groups at the 0.95 confidence level. The highest mean value of released TGF-ß1 (1993.1 pg/mL) was detected in the control group, where the root canal dentin was conditioned with 17% EDTA alone. Regarding the experimental groups, Bio-C Temp released a statistically significantly higher amount of TGF-ß1 (282.14 pg/mL) compared to UltraCal XS (114.28 pg/mL; p = 0.0158). Bio-C Temp affected the release of growth factors from root canal dentin less than UltraCal XS and may therefore serve as an intracanal medicament for regenerative endodontic procedures.

Ameliorative Potential of Bone Marrow-Derived Mesenchymal Stem Cells Versus Prednisolone in a Rat Model of Lung Fibrosis: A Histological, Immunohistochemical, and Biochemical Study.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease of unknown origin with limited treatment options and poor prognosis. The encouraging findings from preclinical investigations utilizing mesenchymal stem cells (MSCs) indicated that they could serve as a promising therapeutic alternative for managing chronic lung conditions, such as IPF. The objective of this study was to compare the efficiency of bone marrow-derived MSCs (BM-MSCs) versus prednisolone, the standard anti-inflammatory medication, in rats with bleomycin (BLM)-induced lung fibrosis. Four groups were created: a control group, a BLM group, a prednisolone-treated group, and a BM-MSCs-treated group. To induce lung fibrosis, 5 mg/kg of BLM was administered intratracheally. BLM significantly increased serum levels of pro-inflammatory cytokines and oxidative stress markers. The disturbed lung structure was also revealed by light and transmission electron microscopic studies. Upregulation in the immune expression of alpha-smooth muscle actin, transforming growth factor beta-1, and Bax was demonstrated. Interestingly, all findings significantly regressed on treatment with prednisolone and BM-MSCs. However, treatment with BM-MSCs showed better results than with prednisolone. In conclusion, BM-MSCs could be a promising approach for managing lung fibrosis.