Electroacupuncture improves myocardial fibrosis in heart failure rats by attenuating ECM collagen deposition through modulation of TGF-β1/Smads signaling pathway

Background: To explore the effects of electroacupuncture on cardiac function and myocardial fibrosis in rat models of heart failure, and to elucidate the underlying mechanism of electroacupuncture in heart failure treatment. Methods: Healthy male Sprague-Dawley rats were allocated into three groups: Sham group, Model group, and electroacupuncture (Model + EA) group, with each group comprising 8 rats. The model underwent a procedure involving the ligation of the left anterior descending coronary artery to induce a model of heart failure. The Model + EA group was used for 7 consecutive days for electroacupuncture of bilateral Shenmen (HT7) and Tongli (HT5), once a day for 30 min each time. Left ventricular parameters in rats were assessed using a small-animal ultrasound machine to analyze changes in left ventricular end-diastolic volume, left ventricular end-systolic volume, left ventricular ejection fraction, and left ventricular fractional shortening. Serum interleukin-1β (IL-1β), cardiac troponin (cTn), and N-terminal brain natriuretic peptide precursor levels were measured using ELISA. Histopathological changes in rat myocardium were observed through HE staining, while collagen deposition in rat myocardial tissue was assessed using the Masson staining method. Picro sirius red staining, immunohistochemical staining, and RT-qPCR were utilized to distinguish between the various types of collagen deposition. The expression level of TGF-β1 and SMAD2/3/4/7 mRNA in rat myocardial tissues was determined using RT-qPCR. Additionally, western blot analysis was conducted to assess the protein expression levels of TGF-β1, SMAD3/7, and p-SMAD3 in rat myocardial tissues. Results: Compared with the Sham group, the left ventricular ejection fraction and left ventricular fractional shortening values of the Model group were significantly decreased (P < 0.01);the left ventricular end-diastolic volume and left ventricular end-systolic volume values were remarkably increased (P < 0.01);serum N-terminal brain natriuretic peptide precursor content was increased (P < 0.01);serum IL-1β and cTn levels were increased (P < 0.01);myocardial collagen volume fraction were increased (P < 0.01);and those of the expression of TGF-β1 and SMAD2/3/4 mRNA was increased (P < 0.01);the expression of SMAD7 mRNA was decreased (P < 0.01);the protein expression levels of TGF-β1, SMAD3, and p-Smad3 were increased (P < 0.01);the protein expression level of SMAD7 was decreased (P < 0.01) in the Model group. Compared to the Model group, the expression levels of the proteins TGF-β1, SMAD3, and p-Smad3 in myocardial tissue were found to be decreased (P < 0.01), and the expression level of the protein SMAD7 was found to be increased (P < 0.01) in the Model + EA group;the collagen volume fraction and deposition of type Ⅰ /Ⅲ collagen were decreased (P < 0.01) in the Model + EA group. Conclusion: Electroacupuncture alleviates myocardial fibrosis in rats with heart failure, and this effect is likely due to attributed to the modulation of the TGF-β1/Smads signaling pathway, which helps reduce collagen deposition in the extracellular matrix.

Piezoelectric film promotes skin wound healing with enhanced collagen deposition and vessels regeneration via upregulation of PI3K/AKT

Skin wounds are common in accidental injuries,and the intricacies of wound repair are closely linked to endogenous electric fields.Electrical stimulation plays a pivotal role in the restorative processes of skin injuries,encompassing collagen deposition,angiogenesis,inflammation,and re-epithelialization.Employing electrical stimulation therapy replicates and enhances the effects of endogenous wound electric fields by applying an external electric field to the wound site,thereby promoting skin wound healing.In this study,we developed a self-powered repetitive mechanical impacts-electrical stimulation(RMI-ES)system utilizing a BaTiO3/polydimethylsiloxane(PDMS)piezoelectric composite film.Compared to conventional electrical stimulation devices,the fabricated piezoelectric composite film efficiently harvests energy from the pressure applied by the stimulation device and the tensile force occurring during natural rat activities.The results demonstrated that piezoelectric stimulation generated by the composite membrane expedited the cell cycle,promoting fibroblast proliferation.Additionally,piezoelectric stimulation induced favorable changes in fibroblast gene expression,including increased expression of transforming growth factor-β1(TGF-β1),connective tissue growth factor(CTGF),collagen 1,collagen 3,vascular endothelial growth factor(VEGF),and alpha-smooth muscle actin(α-SMA),while reducing interleukin-6(IL-6)expression.Transcriptome analysis revealed that piezoelectric stimulation may induce fibroblast migration,proliferation,and collagen expression by influencing PI3K/AKT serine/threonine kinase(AKT)pathways.Further confirmation through the addition of the PI3K inhibitor LY294002 validated that piezoelectric stimulation can regulate the repair process after skin injury through the pathway.Importantly,in vivo results demonstrated that the electric field at the wound site effectively promoted wound healing,reduced inflammation,and stimulated collagen deposition and neovascularization.This study emphasizes the role of the piezoelectric membrane as an effective,safe,and battery-free electrical stimulator crucial for skin wound healing.

Proliferation and tenogenic differentiation of bone marrow mesenchymal stem cells in a porous collagen sponge scaffold

BACKGROUND Collagen is one of the most commonly used natural biomaterials for tendon tissue engineering.One of the possible practical ways to further enhance tendon repair is to combine a porous collagen sponge scaffold with a suitable growth factor or cytokine that has an inherent ability to promote the recruitment,proliferation,and tenogenic differentiation of cells.However,there is an incomplete understanding of which growth factors are sufficient and optimal for the tenogenic differentiation of rat bone marrow mesenchymal stem cells(BMSCs)in a collagen sponge-based 3D culture system.AIM To identify one or more ideal growth factors that benefit the proliferation and tenogenic differentiation of rat BMSCs in a porous collagen sponge scaffold.METHODS We constructed a 3D culture system based on a type I collagen sponge scaffold.The surface topography of the collagen sponge scaffold was observed by scanning electron microscopy.Primary BMSCs were isolated from Sprague-Dawley rats.Cell survival on the surfaces of the scaffolds with different growth factors was assessed by live/dead assay and CCK-8 assay.The mRNA and protein expression levels were confirmed by quantitative real-time polymerase chain reaction and Western blot,respectively.The deposited collagen was assessed by Sirius Red staining.RESULTS Transforming growth factorβ1(TGF-β1)showed great promise in the tenogenic differentiation of BMSCs compared to growth differentiation factor 7(GDF-7)and insulin-like growth factor 1(IGF-1)in both the 2D and 3D cultures,and the 3D culture enhanced the differentiation of BMSCs into tenocytes well beyond the level of induction in the 2D culture after TGF-β1 treatment.In the 2D culture,the proliferation of the BMSCs showed no significant changes compared to the control group after TGF-β1,IGF-1,or GDF-7 treatment.However,TGF-β1 and GDF-7 could increase the cell proliferation in the 3D culture.Strangely,we also found more dead cells in the BMSC-collagen sponge constructs that were treated with TGF-β1.Moreover,TGF-β1 promoted more collagen deposition in both the 2D and 3D cultures.CONCLUSION Collagen sponge-based 3D culture with TGF-β1 enhances the responsiveness of the proliferation and tenogenic differentiation of rat BMSCs.

Inhibition of TGF-β1 by eNOS gene transfer provides cardiac protection after myocardial infarction

Objective： Endothelial nitric oxide synthase （eNOS） and nitric oxide （NO） have been implicated in protection against myocardial ischemia injury. This study was designed to explore a new method of therapy for myocardial injury by eNOS gene transfection. Methods： A rat model of myocardial infarction （MI） was established by left anterior descending （LAD） coronary artery ligation, eNOS gene in an adenovirus vector was delivered locally into the rat heart and hemodynamic parameters were examined after 3 weeks, Matrix metalloproteinase-2 and 9 （MMP-2, MMP-9） mRNA were measured by reverse transcription polymerase chain reaction （RT-PCR）, and the protein levels of eNOS, caspase-3, and transforming grouth factor 131 （TGF-131） were determined by western blot assay. Results： eNOS gene transfer significantly reduced cardiomyocyte apoptosis and improved cardiac function. In addition, eNOS significantly reduced the mRNA levels of MMP-2 and MMP-9. In the eNOS gene transfected group, the activation of caspase-3 and TGF-β1 were decreased. However, the protection was reversed by administration of the NOS inhibitor, N（o））-nitro-l-arginine methyl ester （L-NAME）. Conclusion： These results demonstrate that the eNOS provides cardiac protection after myocardial infarction injury through inhibition of cardiac apoptosis and collagen deposition, and suppression of TGF-β1.

A novel chronic nerve compression model in the rat

Current animal models of chronic peripheral nerve compression are mainly silicone tube models. However, the cross section of the rat sciatic nerve is not a perfect circle, and there are differences in the diameter of the sciatic nerve due to individual differences. The use of a silicone tube with a uniform internal diameter may not provide a reliable and consistent model. We have established a chronic sciatic nerve compression model that can induce demyelination of the sciatic nerve and lead to atrophy of skeletal muscle. In 3-week-old pups and adult rats, the sciatic nerve of the right hind limb was exposed, and a piece of surgical latex glove was gently placed under the nerve. N-butyl-cyanoacrylate was then placed over the nerve, and after it had set, another piece of glove latex was placed on top of the target area and allowed to adhere to the first piece to form a sandwich-like complex. Thus, a chronic sciatic nerve compression model was produced. Control pups with latex or N-butyl-cyanoacrylate were also prepared. Functional changes to nerves were assessed using the hot plate test and electromyography. Immunofluorescence and electron microscopy analyses of the nerves were performed to quantify the degree of neuropathological change. Masson staining was conducted to assess the degree of fibrosis in the gastrocnemius and intrinsic paw muscles. The pup group rats subjected to nerve compression displayed thermal hypoesthesia and a gradual decrease in nerve conduction velocity at 2 weeks after surgery. Neuropathological studies demonstrated that the model caused nerve demyelination and axonal irregularities and triggered collagen deposition in the epineurium and perineurium of the affected nerve at 8 weeks after surgery. The degree of fibrosis in the gastrocnemius and intrinsic paw muscles was significantly increased at 20 weeks after surgery. In conclusion, our novel model can reproduce the functional and histological changes of chronic nerve compression injury that occurs in humans and it will be a useful new tool for investigating the mechanisms underlying chronic nerve compression.

Platelet-rich plasma accelerates skin wound healing by promoting re-epithelialization

Background:Autologous platelet-rich plasma(PRP)has been suggested to be effective for wound healing.However,evidence for its use in patients with acute and chronic wounds remains insufficient.The aims of this study were to comprehensively examine the effectiveness,synergy and possible mechanism of PRP-mediated improvement of acute skin wound repair.Methods:Full-thickness wounds were made on the back of C57/BL6 mice.PRP or saline solution as a control was administered to the wound area.Wound healing rate,local inflammation,angiogenesis,re-epithelialization and collagen deposition were measured at days 3,5,7 and 14 after skin injury.The biological character of epidermal stem cells(ESCs),which reflect the potential for re-epithelialization,was further evaluated in vitro and in vivo.Results:PRP strongly improved skin wound healing,which was associated with regulation of local inflammation,enhancement of angiogenesis and re-epithelialization.PRP treatment significantly reduced the production of inflammatory cytokines interleukin-17A and interleukin-1β.An increase in the local vessel intensity and enhancement of re-epithelialization were also observed in animals with PRP administration and were associated with enhanced secretion of growth factors such as vascular endothelial growth factor and insulin-like growth factor-1.Moreover,PRP treatment ameliorated the survival and activated the migration and proliferation of primary cultured ESCs,and these effects were accompanied by the differentiation of ESCs into adult cells following the changes of CD49f and keratin 10 and keratin 14.Conclusion:PRP improved skin wound healing by modulating inflammation and increasing angiogenesis and re-epithelialization.However,the underlying regulatory mechanism needs to be investigated in the future.Our data provide a preliminary theoretical foundation for the clinical administration of PRP in wound healing and skin regeneration.

P311 promotes typeⅡtransforming growth factor-βreceptor mediated fibroblast activation and granulation tissue formation in wound healing

Background:P311,a highly conserved 8 kDa intracellular protein,has recently been reported to play an important role in aggravating hypertrophic scaring by promoting the differentiation and secretion of fibroblasts.Nevertheless,how P311 regulates the differentiation and function of fibroblasts to affect granulation tissue formation remains unclear.In this work,we studied the underlying mechanisms via which P311 affects fibroblasts and promotes acute skin wound repair.Methods:To explore the role of P311,both in vitro and in vivo wound-healing models were used.Full-thickness skin excisional wounds were made in wild-type and P311−/−C57 adult mice.Wound healing rate,re-epithelialization,granulation tissue formation and collagen deposition were measured at days 3,6 and 9 after skin injury.The biological phenotypes of fibroblasts,the expression of target proteins and relevant signaling pathways were examined both in vitro and in vivo.Results:P311 could promote the proliferation and differentiation of fibroblasts,enhance the ability of myofibroblasts to secrete extracellular matrix and promote cell contraction,and then facilitate the formation of granulation tissue and eventually accelerate skin wound closure.Importantly,we discovered that P311 acts via up-regulating the expression of type II transforming growth factor-βreceptor(TGF-βRII)in fibroblasts and promoting the activation of the TGF-βRII-Smad signaling pathway.Mechanistically,the mammalian target of rapamycin signaling pathway is closely implicated in the regulation of the TGF-βRII-Smad pathway in fibroblasts mediated by P311.Conclusions:P311 plays a critical role in activation of the TGF-βRII-Smad pathway to promote fibroblast proliferation and differentiation as well as granulation tissue formation in the process of skin wound repair.