PAPP-A enhances the antioxidative effects of IGF-1 during bovine in vitro embryo production.

We investigated whether exogenous pregnancy-associated plasma protein-A (PAPP-A) enhances the antioxidant role of insulin-like growth factor-1 (IGF-1) in bovine in vitro embryo production (IVP). We performed standard in vitro maturation (IVM) and in vitro culture (IVC) or added menadione to promote an oxidative stressed microenvironment and evaluated the antioxidant effect of IGF-1 alone or in combination with PAPP-A (IGF-1/PAPP-A). In IVM, the treatments did not affect oocyte nuclear development, total GSH content, cumulus cell gene expression, and blastocyst yield. Nevertheless, IGF-1/PAPP-A treatment prevented an increase in reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) levels. In IVC, the treatments did not affect the total GSH content on blastocysts and IVC media, but IGF-1 and IGF-1/PAPP-A treatments increased blastocyst yield compared to the menadione group. In addition, IGF-1/PAPP-A treatment had lower ROS levels and regulated genes related to embryonic quality compared to the control and menadione groups. Overall, we showed that PAPP-A could enhance the antioxidant role of IGF-1 during IVP in cattle by avoiding higher ROS levels in oocytes and blastocysts and modulating the transcriptional abundance of genes involved in oxidative protection and embryonic quality.

Insulin promotes the bone formation capability of human dental pulp stem cells through attenuating the IIS/PI3K/AKT/mTOR pathway axis.

BACKGROUND: Insulin has been known to regulate bone metabolism, yet its specific molecular mechanisms during the proliferation and osteogenic differentiation of dental pulp stem cells (DPSCs) remain poorly understood. This study aimed to explore the effects of insulin on the bone formation capability of human DPSCs and to elucidate the underlying mechanisms. METHODS: Cell proliferation was assessed using a CCK-8 assay. Cell phenotype was analyzed by flow cytometry. Colony-forming unit-fibroblast ability and multilineage differentiation potential were evaluated using Toluidine blue, Oil red O, Alizarin red, and Alcian blue staining. Gene and protein expressions were quantified by real-time quantitative polymerase chain reaction and Western blotting, respectively. Bone metabolism and biochemical markers were analyzed using electrochemical luminescence and chemical colorimetry. Cell adhesion and growth on nano-hydroxyapatite/collagen (nHAC) were observed with a scanning electron microscope. Bone regeneration was assessed using micro-CT, fluorescent labeling, immunohistochemical and hematoxylin and eosin staining. RESULTS: Insulin enhanced the proliferation of human DPSCs as well as promoted mineralized matrix formation in a concentration-dependent manner. 10- 6 M insulin significantly up-regulated osteogenic differentiation-related genes and proteins markedly increased the secretion of bone metabolism and biochemical markers, and obviously stimulated mineralized matrix formation. However, it also significantly inhibited the expression of genes and proteins of receptors and receptor substrates associated with insulin/insulin-like growth factor-1 signaling (IIS) pathway, obviously reduced the expression of the phosphorylated PI3K and the ratios of the phosphorylated PI3K/total PI3K, and notably increased the expression of the total PI3K, phosphorylated AKT, total AKT and mTOR. The inhibitor LY294002 attenuated the responsiveness of 10- 6 M insulin to IIS/PI3K/AKT/mTOR pathway axis, suppressing the promoting effect of insulin on cell proliferation, osteogenic differentiation and bone formation. Implantation of 10- 6 M insulin treated DPSCs into the backs of severe combined immunodeficient mice and the rabbit jawbone defects resulted in enhanced bone formation. CONCLUSIONS: Insulin induces insulin resistance in human DPSCs and effectively promotes their proliferation, osteogenic differentiation and bone formation capability through gradually inducing the down-regulation of IIS/PI3K/AKT/mTOR pathway axis under insulin resistant states.

A triple growth factor strategy for optimizing bone augmentation in mice.

With dental implant treatment becoming the gold standard, the need for effective bone augmentation prior to implantation has grown. This study aims to evaluate a bone augmentation strategy integrating three key growth factors: bone morphogenetic protein-2 (BMP-2), insulin-like growth factor 1 (IGF-1), and vascular endothelial growth factor (VEGF). Collagen scaffolds incorporating BMP-2, IGF-1, or VEGF were fabricated and categorized into five groups based on their content: scaffold alone; BMP-2 alone (BMP-2); BMP-2 and IGF-1 (BI); BMP-2, IGF-1, and VEGF (BIV); and BMP-2 and IGF-1 with an earlier release of VEGF (BI + V). The prepared scaffolds were surgically implanted into the calvarias of C57BL/6JJcl mice, and hard tissue formation was assessed after 10 and 28 days through histological, tomographic, and biochemical analyses. The combination of BMP-2 and IGF-1 induced a greater volume of hard tissue augmentation compared with that of BMP-2 alone, regardless of VEGF supplementation, and these groups had increased levels of cartilage compared with others. The volume of hard tissue formation was greatest in the BIV group. In contrast, the BI + V group exhibited a hard tissue volume similar to that of the BI group. While VEGF and CD31 levels were highest in the BIV group at 10 days, there was no correlation at the same time point between hard tissue formation and the quantity of M2 macrophages. In conclusion, the simultaneous release of BMP-2, IGF-1, and VEGF proved to be effective in promoting bone augmentation.

GLP-2 ameliorates D-galactose induced muscle aging by IGF-1/Pi3k/Akt/FoxO3a signaling pathway in C2C12 cells and mice.

BACKGROUND: The study aimed to investigate the effect of Glucagon-like peptide-2 (GLP-2) on muscle aging in vivo and in vitro. METHODS: Six-week-old C57BL/6J mice were administered with D-galactose (200 mg/kg/day, intraperitoneally) for 8weeks, followed by daily subcutaneous injections of GLP-2 (300 or 600 µg/kg/day) for 4weeks. Skeletal muscle function and mass were evaluated using relative grip strength and muscle weight. The sizes and types of muscle fibers and apoptosis were assessed through histological analysis, immunofluorescence staining, and TUNEL staining, respectively. C2C12 myotubes were treated with D-galactose (40 mg/mL) and GLP-2. Protein expression of differentiation-related myogenic differentiation factor D (MyoD), myogenin (MyoG), and myosin heavy chain (Myhc), degradation-related Muscle RING finger 1 (MuRF-1), and muscle atrophy F-box (MAFbx)/Atrogin-1, and apoptosis-related B-cell leukemia/lymphoma 2 (Bcl-2) and Bax, were assessed using western blots. The Pi3k inhibitor LY294002 was applied to investigate whether GLP-2 regulated myogenesis and myotube aging via IGF-1/Pi3k/Akt/FoxO3a signaling pathway. RESULTS: The results demonstrated that GLP-2 significantly reversed the decline in muscles weight, relative grip strength, diameter, and cross-sectional area of muscle fibers induced by D-galactose in mice. Apart from suppressing the expressions of MuRF-1 and Atrogin-1 in the muscles and C2C12 myotubes, GLP-2 significantly increased the expressions of MyoD, MyoG, and Myhc compared to the D-galactose. GLP-2 significantly suppressed cell apoptosis. Western blot analysis indicated that the regulation of GLP-2 may be attributed to the activation of theIGF-1/Pi3k/Akt/FoxO3a phosphorylation pathway. CONCLUSIONS: This study suggested that GLP-2 ameliorated D-galactose induced muscle aging by IGF-1/Pi3k/Akt/FoxO3a pathway.

Injectable thermo-responsive Poloxamer hydrogel/methacrylate gelatin microgels stimulates bone regeneration through biomimetic programmed release of SDF-1a and IGF-1.

Injectable hydrogels, offering adaptable drug delivery of growth factors (GFs), hold promise for treating bone defects. To optimize osteogenic efficacy, the release of GFs should mirror the natural bone healing. We developed an injectable thermo-responsive hydrogel/microgels platform for dual GF delivery for bone regeneration. Stromal cell-derived factor-1 alpha (SDF-1a) and the Methacrylate Gelatin (GelMA) microgels which encapsulated insulin-like growth factor-1 (IGF-1) loaded liposomes (Ls) were introduced into Poloxamer 407 (P407) hydrogel matrix. This system achieved the biomimetic release profile of SDF-1a and IGF-1, which covered the early stage from day 1 to 7 and the continuous stage from day 5 to 21, respectively. In vitro study confirmed the enhanced migration, osteogenic biomarker expression, and matrix mineralization of the bone marrow mesenchymal stem cells (BMSCs) co-cultivated with the dual GFs delivering hydrogel/microgels. Transcriptome sequencing revealed that the potential mechanism was associated with mitogen-activated protein kinase (MAPK) signaling activation and its downstream ribosomal protein S6 kinase 2 (RSK2) upregulation. In a critical-sized calvarial defect model in Sprague-Dawley (SD) rats, the injectable hydrogel/microgels system promoted significant bone regeneration. Collectively, our study suggested the current hydrogel/microgels system with the biomimetic release of SDF-1a and IGF-1 efficiently promoted bone regeneration, informing the future development of GF delivery systems intended for bone regeneration therapies.

The involvement of Akt, mTOR, and S6K in the in vivo effect of IGF-1 on the regulation of rat cardiac Na+/K+-ATPase.

BACKGROUND: We previously demonstrated that insulin-like growth factor-1 (IGF-1) regulates sodium/potassium adenosine triphosphatase (Na+/K+-ATPase) in vascular smooth muscle cells (VSMC) via phosphatidylinositol-3 kinase (PI3K). Taking into account that others' work show that IGF-1 activates the PI3K/protein kinase B (Akt) signaling pathway in many different cells, we here further questioned if the Akt/mammalian target of rapamycin (mTOR)/ribosomal protein p70 S6 kinase (S6K) pathway stimulates Na+/K+-ATPase, an essential protein for maintaining normal heart function. METHODS AND RESULTS: There were 14 adult male Wistar rats, half of whom received bolus injections of IGF-1 (50 µg/kg) for 24 h. We evaluated cardiac Na+/K+-ATPase expression, activity, and serum IGF-1 levels. Additionally, we examined the phosphorylated forms of the following proteins: insulin receptor substrate (IRS), phosphoinositide-dependent kinase-1 (PDK-1), Akt, mTOR, S6K, and α subunit of Na+/K+-ATPase. Additionally, the mRNA expression of the Na+/K+-ATPase α1 subunit was evaluated. Treatment with IGF-1 increases levels of serum IGF-1 and stimulates Na+/K+-ATPase activity, phosphorylation of α subunit of Na+/K+-ATPase on Ser23, and protein expression of α2 subunit. Furthermore, IGF-1 treatment increased phosphorylation of IRS-1 on Tyr1222, Akt on Ser473, PDK-1 on Ser241, mTOR on Ser2481 and Ser2448, and S6K on Thr421/Ser424. The concentration of IGF-1 in serum positively correlates with Na+/K+-ATPase activity and the phosphorylated form of mTOR (Ser2448), while Na+/K+-ATPase activity positively correlates with the phosphorylated form of IRS-1 (Tyr1222) and mTOR (Ser2448). CONCLUSION: These results indicate that the Akt/mTOR/S6K signalling pathway may be involved in the IGF-1 regulating cardiac Na+/K+-ATPase expression and activity.

Impact of media supplements FGF2, LIF and IGF1 on the genome activity of porcine embryos produced in vitro.

In this article, we focused on the impact of precisely chemically modified FLI maturation medium enriched with fibroblast growth factor 2 (FGF2), leukemia inhibitory factor (LIF), insulin-like growth factor 1 (IGF1), and polyvinyl alcohol (PVA) and its potential to improve the efficiency of in vitro production of porcine embryos. We hypothesized that enhancing the composition of the maturation medium could result in an elevated production of embryos in vitro and can affect EGA. FLI medium resulted in a significantly higher rate of oocyte blastocyst maturation and formation compared to the control DMEM medium. In addition, immunocytochemical labelling confirmed the detection of UBF in 4-cell FLI parthenogenic embryos, suggesting similarities with natural embryo development. Through RNAseq analysis, upregulated genes present in 4-cell FLI embryos were found to play key roles in important biological processes such as cell proliferation, cell differentiation, and transcriptional regulation. Based on our findings, we demonstrated the positive influence of FLI medium in the evaluation of in vitro embryo production, EGA detection, transcriptomic and proteomic profile, which was confirmed by the positive activation of the embryonal genome in the 4-cell stage of parthenogenetically activated embryos.

IGF-1R mediates crosstalk between nasopharyngeal carcinoma cells and osteoclasts and promotes tumor bone metastasis.

BACKGROUND: Nasopharyngeal carcinoma (NPC) poses a significant health burden in specific regions of Asia, and some of NPC patients have bone metastases at the time of initial diagnosis. Bone metastasis can cause pathologic fractures and pain, reducing patients' quality of life, and is associated with worse survival. This study aims to unravel the complex role of insulin-like growth factor 1 receptor (IGF-1R) in NPC bone metastasis, offering insights into potential therapeutic targets. METHODS: We assessed IGF-1R expression in NPC cells and explored its correlation with bone metastasis. Experiments investigated the impact of osteoclast-secreted IGF-1 on the IGF-1R/AKT/S6 pathway in promoting NPC cell proliferation within the bone marrow. Additionally, the reciprocal influence of tumor-secreted Granulocyte-macrophage colony-stimulating factor (GM-CSF) on osteoclast differentiation and bone resorption was examined. The effects of IGF-1 neutralizing antibody, IGF-1R specific inhibitor (NVP-AEW541) and mTORC inhibitor (rapamycin) on nasopharyngeal carcinoma bone metastasis were also explored in animal experiments. RESULTS: Elevated IGF-1R expression in NPC cells correlated with an increased tendency for bone metastasis. IGF-1, secreted by osteoclasts, activated the IGF-1R/AKT/S6 pathway, promoting NPC cell proliferation in the bone marrow. Tumor-secreted GM-CSF further stimulated osteoclast differentiation, exacerbating bone resorption. The IGF-1 neutralizing antibody, NVP-AEW541 and rapamycin were respectively effective in slowing down the rate of bone metastasis and reducing bone destruction. CONCLUSION: The intricate interplay among IGF-1R, IGF-1, and GM-CSF highlights potential therapeutic targets for precise control of NPC bone metastasis, providing valuable insights for developing targeted interventions.

In Vitro Assessment of Bacterial Adhesion and Biofilm Formation on Novel Bioactive, Biodegradable Electrospun Fiber Meshes Intended to Support Tendon Rupture Repair.

The surgical repair of a ruptured tendon faces two major problems: specifically increased fibrous adhesion to the surrounding tissue and inferior mechanical properties of the scar tissue compared to the native tissue. Bacterial attachment to implant materials is an additional problem as it might lead to severe infections and impaired recovery. To counteract adhesion formation, two novel implant materials were fabricated by electrospinning, namely, a random fiber mesh containing hyaluronic acid (HA) and poly(ethylene oxide) (PEO) in a ratio of 1:1 (HA/PEO 1:1) and 1:4 (HA/PEO 1:4), respectively. Electrospun DegraPol (DP) treated with silver nanoparticles (DP-Ag) was developed to counteract the bacterial attachment. The three novel materials were compared to the previously described DP and DP with incorporated insulin-like growth factor-1 (DP-IGF-1), two implant materials that were also designed to improve tendon repair. To test whether the materials are prone to bacterial adhesion and biofilm formation, we assessed 10 strains of Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Enterococcus faecalis, known for causing nosocomial infections. Fiber diameter, pore size, and water contact angle, reflecting different degrees of hydrophobicity, were used to characterize all materials. Generally, we observed higher biofilm formation on the more hydrophobic DP as compared to the more hydrophilic DP-IGF-1 and a trend toward reduced biofilm formation for DP treated with silver nanoparticles. For the two HA/PEO implants, a similar biofilm formation was observed. All tested materials were highly prone to bacterial adherence and biofilm formation, pointing toward the need of further material development, including the optimized incorporation of antibacterial agents such as silver nanoparticles or antibiotics.

IGF-PLGA microspheres promote angiogenesis and accelerate skin flap repair and healing by inhibiting oxidative stress and regulating the Ang 1/Tie 2 signaling pathway.

Random flaps are widely used in the treatment of injuries, tumors, congenital malformations, and other diseases. However, postoperative skin flaps are prone to ischemic necrosis, leading to surgical failure. Insulin-like growth factor- 1(IGF-1) belongs to the IGF family and exerts its growth-promoting effects in various tissues through autocrine or paracrine mechanisms. Its application in skin flaps and other traumatic diseases is relatively limited. Poly (lactic-co-glycolic acid) (PLGA) is a degradable high-molecular-weight organic compound commonly used in biomaterials. This study prepared IGF-PLGA sustained-release microspheres to explore their impact on the survival rate of flaps both in vitro and in vivo, as well as the mechanisms involved. The research results demonstrate that IGF-PLGA has a good sustained-release effect. At the cellular level, it can promote 3T3 cell proliferation by inhibiting oxidative stress, inhibit apoptosis, and enhance the tube formation ability of human umbilical vein endothelial cells (HUVEC) . At the animal level, it accelerates flap healing by promoting vascularization through the inhibition of oxidative stress. Furthermore, this study reveals the role of IGF-PLGA in activating the Angiopoietin-1(Ang1)/Tie2 signaling pathway in promoting flap vascularization, providing a strong theoretical basis and therapeutic target for the application of IGF-1 in flaps and other traumatic diseases.

Effects of exogenous steroid hormones on growth, body color, and gonadal development in the Opsariichthys bidens.

To investigate the effects of exogenous steroid hormones on growth, body color, and gonadal development in the Opsariichthys bidens (O. bidens), synthetic methyltestosterone (MT) and 17ß-estradiol (E2) were used for 28 days' treatment of 4-month-old O. bidens before the breeding season. Our results suggested that MT had a significant growth-promoting effect (P < 0.05), whereas E2 played an inhibitory role. On the body surface, the females in the MT group showed gray stripes, and the fish in other groups showed no obvious stripes. The males with MT treatment displayed brighter blue-green stripes compared to the CK and E2 groups. The histological analysis showed that the MT significantly promoted testes development in males, blocked oocyte development, and caused massive apoptosis in females, whereas the E2 group promoted ovarian development and inhibited testes development. Based on qRT-PCR analysis, in females, the expression of igf-1, dmrt1, and cyp19a1a genes revealed that E2 treatment resulted in down-regulation of igf-1 expression and up-regulation of cyp19a1a expression. In males, igf-1 and dmrt1 were significantly up-regulated after MT treatment, and E2 treatment led to down-regulation of igf-1. Therefore, this study demonstrates that MT and E2 play an important role in reversing the morphological sex characteristics of females and males.

Accelerative effects of alginate-chitosan/titanium oxide@geraniol nanosphere hydrogels on the healing process of wounds infected with Acinetobacter baumannii and Streptococcus pyogenes bacteria.

This study was conducted to evaluate the effects of alginate-chitosan/titanium oxide/geraniol (Alg-Csn/TiO2@GRL nanosphere) nanospheres hydrogels on the healing process of the wounds infected with Acinetobacter baumannii and Streptococcus pyogenes bacteria. The nanospheres were successfully synthesized and their physicochemical properties such as DLS, FTIR, FE-SEM, TEM, XRD and also their safety and in-vitro antibacterial activity were assessed and confirmed. Following induction of the infected wounds, the mice were treated with s base ointment (Control), mupirocin® as standard control group and also hydrogels prepared from Alg-Csn@GRL, Alg-Csn/TiO2 and Alg-Csn/TiO2@GRL. Wound contraction, total bacterial count, expression of bFGF, VEGF, IGF-1, CD68 and COL-1 A, iNOS and eNOS were measured. The results showed the treatment of wounds with Alg-Csn/TiO2@GRL hydrogels significantly accelerated wound contraction, decreased total bacterial count and reduced the expressions of CD68, iNOS and eNOS and increased the expressions of VEGF, bFGF, IGF-1 and COL-1 A compared with other groups. It can be concluded that Alg-Csn/TiO2@GRL hydrogels expedite the wound healing process by their effects on bacteria and subsequently inflammation and increasing the expression of proliferative genes. The Alg-Csn/TiO2@GRL hydrogel can be utilized in combination with other agents for the treatment of infected wounds after future clinical studies.

Microvascular smooth muscle cells exhibit divergent phenotypic switching responses to platelet-derived growth factor and insulin-like growth factor 1.

OBJECTIVE: Vascular smooth muscle cell (VSMC) phenotypic switching is critical for normal vessel formation, vascular stability, and healthy brain aging. Phenotypic switching is regulated by mediators including platelet derived growth factor (PDGF)-BB, insulin-like growth factor (IGF-1), as well as transforming growth factor-ß (TGF-ß) and endothelin-1 (ET-1), but much about the role of these factors in microvascular VSMCs remains unclear. METHODS: We used primary rat microvascular VSMCs to explore PDGF-BB- and IGF-1-induced phenotypic switching. RESULTS: PDGF-BB induced an early proliferative response, followed by formation of polarized leader cells and rapid, directionally coordinated migration. In contrast, IGF-1 induced cell hypertrophy, and only a small degree of migration by unpolarized cells. TGF-ß and ET-1 selectively inhibit PDGF-BB-induced VSMC migration primarily by repressing migratory polarization and formation of leader cells. Contractile genes were downregulated by both growth factors, while other genes were differentially regulated by PDGF-BB and IGF-1. CONCLUSIONS: These studies indicate that PDGF-BB and IGF-1 stimulate different types of microvascular VSMC phenotypic switching characterized by different modes of cell migration. Our studies are consistent with a chronic vasoprotective role for IGF-1 in VSMCs in the microvasculature while PDGF is more involved in VSMC proliferation and migration in response to acute activities such as neovascularization. Better understanding of the nuances of the phenotypic switching induced by these growth factors is important for our understanding of a variety of microvascular diseases.

Establishment and Characterization of Amitrole-Induced Mouse Thyroid Adenomatous Nodule-Derived Cell Lines.

Background: Thyroid cancer cell lines have been of great value for the study of thyroid cancer. However, the availability of benign thyroid adenoma cell lines is limited. Methods: Cell lines were established from thyroid adenomatous nodules that developed in mice treated with the goitrogen amitrole. Expression of epithelial, mesenchymal, and thyroid markers of these established cell lines was determined, and the effect of lentivirus-transduced overexpression of NKX2-1, a master regulator of thyroid development, on the thyroid marker expression was examined. Signal transduction and cell proliferation were evaluated after treatment with insulin-like growth factor-I (IGF-I) and the selective IGF-I receptor (IGF-IR) inhibitor NVP-ADW742. Xenograft studies were performed to examine tumorigenicity of the cells in mice. Whole-genome sequencing (WGS) was used to comprehensively determine the genetic mutations in the established two cell lines. Results: Five mouse thyroid adenomatous nodules-derived cell lines named CAT (cells from amitrole-treated thyroids) were established. Among these, two cell lines, CAT458/458s (CAT458s: a subline of CAT458) and CAT459, were found to be positive for epithelial markers and negative for a mesenchymal marker. NKX2-1-positive CAT459 cells showed higher messenger RNA (mRNA) expression of some thyroid differentiation markers than NKX2-1-negative CAT458s cells, and NKX2-1 overexpression increased and/or induced their expression. IGF-I signaling was transduced in thyrotropin receptor (Tshr)-negative CAT458s and 459 cells, and NVP-ADW742 suppressed their proliferation. No tumors developed in mice after subcutaneous injection of CAT458s or 459 cells. The WGS analysis revealed the presence of missense mutations in the tumor suppressor genes such as Polk (encoding DNA polymerase kappa) and Tgfb1 (encoding transforming growth factor beta 1), while no mutations were found in the prominent thyroid cancer-related genes Braf, Trp53 (encoding p53), and Tert (encoding telomerase reverse transcriptase). Conclusions: Two mouse thyroid adenomatous nodule-derived cell lines with different thyroid differentiation marker expression were established. NKX2-1 induced partial differentiation of these cell lines. They lacked tumorigenicity and prominent gene mutations involved in thyroid cancer development, while missense mutations were found in some tumor suppressors as revealed by WGS. The CAT458s and 459 provide a new tool to further clarify the process of thyroid multistep carcinogenesis and differentiation.

[Effect and mechanism of Zuogui Pills on neural function recovery in ischemic stroke mice based on OPN/IGF-1/mTOR].

To explore the effect and mechanism of Zuogui Pills in promoting neural tissue recovery and functional recovery in mice with ischemic stroke. Male C57BL/6J mice were randomly divided into a sham group, a model group, and low-, medium, and high-dose Zuogui Pills groups(3.5, 7, and 14 g·kg~(-1)), with 15 mice in each group. The ischemic stroke model was established using photochemical embolization. Stiker remove and irregular ladder walking behavioral tests were conducted before modeling and on days 7, 14, 21, and 28 after medication. Triphenyl tetrazolium chloride(TTC) staining was performed on day 3 after modeling, and T2-weighted imaging(T2WI) and diffusion-weighted imaging(DWI) were performed on day 28 after medication to evaluate the extent of brain injury. Hematoxylin-eosin(HE) staining was performed to observe the histology of the cerebral cortex. Axonal marker proteins myelin basic protein(MBP), growth-associated protein 43(GAP43), mammalian target of rapamycin(mTOR), and its downstream phosphorylated s6 ribosomal protein(p-S6), as well as mechanism-related proteins osteopontin(OPN) and insulin-like growth factor 1(IGF-1), were detected using immunofluorescence and Western blot. Zuogui Pills had a certain restorative effect on the neural function impairment caused by ischemic stroke in mice. TTC staining showed white infarct foci in the sensory-motor cortex area, and T2WI imaging revealed cystic necrosis in the sensory-motor cortex area. The Zuogui Pills groups showed less brain tissue damage, fewer scars, and more capillaries. The number of neuronal axons in those groups was higher than that in the model group, and neuronal activity was stronger. The expression of GAP43, OPN, IGF-1, and mTOR proteins in the Zuogui Pills groups was higher than that in the model group. In summary, Zuogui Pills can promote the recovery of neural function and axonal growth in mice with ischemic stroke, and its mechanism may be related to the activation of the OPN/IGF-1/mTOR signaling pathway.

Electrospun PCL Nerve Conduit Filled with GelMA Gel for CNTF and IGF-1 Delivery in Promoting Sciatic Nerve Regeneration in Rat.

Neural tissue engineering is an essential strategy to repair long-segment peripheral nerve defects. Modification of the nerve conduit is an effective way to improve the local microenvironment of the injury site and facilitate nerve regeneration. However, the concurrent release of multiple growth cues that regulate the activity of Schwann cells and neurons remains a challenge. The present study involved the fabrication of a composite hydrogel, specifically methacrylate-anhydride gelatin-ciliary neurotrophic factor/insulin-like growth factor-1 (GelMA-CNTF/IGF-1), with the aim of providing a sustained release of CNTF and IGF-1. The GelMA-CNTF/IGF-1 hydrogels exhibited a swelling rate of 10.2% following a 24 h incubation in vitro. In vitro, GelMA hydrogels demonstrated a high degree of efficiency in the sustained release of CNTF and IGF-1 proteins, with a release rate of 85.9% for CNTF and 90.9% for IGF-1 shown at day 28. In addition, the GelMA-CNTF/IGF-1 composite hydrogel promoted the proliferation of Schwann cells and the production of nerve growth factor (NGF), connective tissue growth factor (CTGF), fibronectin, and laminin and also considerably promoted the axonal growth of neurons. Furthermore, GelMA-CNTF/IGF-1 hydrogels were loaded into PCL electrospun nerve conduits to repair 15 mm sciatic nerve defects in rats. In vivo studies indicated that PCL-GelMA-CNTF/IGF-1 could efficiently accelerate the regeneration of the rat sciatic nerve, promote the formation of the myelin sheath of new axons, promote the electrophysiological function of regenerated nerves, and eventually improve the recovery of motor function in rats. Overall, the PCL-GelMA-CNTF/IGF-1 scaffold presents an attractive new approach for generating an optimal therapeutic alternative for peripheral nerve restoration.

IGF­1 inhibits palmitic acid­induced mitochondrial apoptosis in macrophages.

Insulin growth factor­1 (IGF­1) is an endocrine regulator that plays an important role in normal growth and development. IGF­1 mediated effects may result in protecting macrophages from immunometabolic response. However, it is unclear whether IGF­1 has a protective effect on fatty acid­induced macrophages damage. In the present study, THP­1 cells were differentiated into macrophages and stimulated with palmitic acid (PA) in the absence or presence of IGF­1. Macrophages apoptosis was measured by Cell Counting Kit­8 assay, flow cytometry, Hoechst 33342 staining and western blotting. The mitochondrial damage was evaluated using JC­1 staining and mitochondrial reactive oxygen species detection. The activation of mitophagy was assessed using immunofluorescence and western blotting. As a result, IGF­1 significantly restored the survival rate in macrophages, while the apoptosis was inhibited through mitochondrial pathway. In addition, IGF­1 protected the mitochondrial damage induced by PA. Furthermore, PA induced mitophagy via phosphatase and tensin homolog­induced putative kinase protein 1/Parkin, which was reversed by IGF­1. Taken together, the present study demonstrated the protective effect of IGF­1 on PA­induced mitochondrial apoptosis in macrophages, which might provide a potential therapeutic strategy for treatment of lipotoxicity.

Evaluation of Stem Cell Laden Collagen + Polycaprolactone + Multi-Walled Carbon Nano-Tubes Nano-Neural Scaffold with and Without Insulin Like Growth Factor-I For Sciatic Nerve Regeneration Post Crush Injury in Wistar Rats.

BACKGROUND/AIMS: All body functions are activated, synchronized and controlled by a substantial, complex network, the nervous system. Upon injury, pathophysiology of the nerve injury proceeds through different paths. The axon may undergo a degenerative retraction from the site of injury for a short distance unless the injury is near to the cell body, in which case it continues to the soma and undergoes retrograde neuronal degeneration. Otherwise, the distal section suffers from Wallerian degeneration, which is marked by axonal swelling, spheroids, and cytoskeleton degeneration. The objective of the study was to evaluate the potential of mesenchymal stem cell laden neural scaffold and insulin-like growth factor I (IGF-I) in nerve regeneration following sciatic nerve injury in a rat model. METHODS: The animals were anaesthetized and a cranio-lateral incision over left thigh was made. Sciatic nerve was exposed and crush injury was introduced for 90 seconds using haemostat at second locking position. The muscle and skin were sutured in routine fashion and thus the rat model of sciatic crush injury was prepared. The animal models were equally distributed into 5 different groups namely A, B, C, D and E and treated with phosphate buffer saline (PBS), carbon nanotubes based neural scaffold only, scaffold with IGF-I, stem cell laden scaffold and stem cell laden scaffold with IGF-I respectively. In vitro scaffold testing was performed. The nerve regeneration was assessed based on physico-neuronal, biochemical, histopathological examination, and relative expression of NRP-1, NRP-2 and GAP-43 and scanning electron microscopy. RESULTS: Sciatic nerve injury model with crush injury produced for 90 seconds was standardized and successfully used in this study. All the biochemical parameters were in normal range in all the groups indicating no scaffold related changes. Physico-neuronal, histopathological, relative gene expression and scanning electron microscopy observations revealed appreciable nerve regeneration in groups E and D, followed by C and B. Restricted to no regeneration was observed in group A. CONCLUSION: Carbon nanotubes based scaffold provided electro-conductivity for proper neuronal regeneration while rat bone marrow-derived mesenchymal stem cells were found to induce axonal sprouting, cellular transformation; whereas IGF-I induced stem cell differentiation, myelin synthesis, angiogenesis and muscle differentiation.

Effect of membrane stabilizers on semen quality and sperm membrane protein expression during cryopreservation of goat semen.

BACKGROUND: Semen cryopreservation is a complex process during which there is alteration in the expression of sperm and seminal plasma proteins, molecular weight of protein or loss of membrane proteins during the process. In order to compensate for these changes, different membrane stabilizers are used in freezing semen extenders. However, there is scarcity of such studies during cryopreservation of goat semen. OBJECTIVE: To investigate the effect of membrane stabilizers on sperm membrane protein expression during cryopreservation of goat semen. MATERIALS AND METHODS: A total of 36 semen ejaculates from nine Assam Hill Goat bucks aged 2 to 2.5 years was collected by artificial vagina method. Three membrane stabilizers, each at two different concentrations viz. 50 and 80 mM sucrose, 50 and 100 mM trehalose, and 100 and 150 ng per mL IGF-1 (insulin-like growth factor 1 protein) were added to Tris-citric acid fructose egg yolk glycerol (TCFEYG) extender and semen samples were cryopreserved. The sperm membrane protein profile was studied in fresh and cryopreserved semen by SDS-PAGE. RESULTS: SDS- PAGE of sperm membrane extract of fresh semen revealed the presence of 24 protein bands with molecular weights ranging from 10 kDa to 240 kDa. Samples supplemented with 50 mM sucrose and 80 mM sucrose revealed 21 protein bands with molecular weights ranging from 10 kDa to 240 kDa. All the 21 protein bands were same as those observed in the sperm membrane of fresh spermatozoa, except that the 23 kDa, 29 kDa and 42 kDa bands were absent in frozen semen. Similarly, frozen semen extended with 50 mM trehalose and 100 mM trehalose revealed 22 protein bands with molecular weights ranging from 10 kDa to 240 kDa, but lacking the 29 kDa and 42 kDa bands. Proteins with molecular weights of 29 kDa, 130 kDa and 240 kDa were absent in frozen semen supplemented with 100 ng per mL IGF-1 and 150 ng per mL IGF-1. CONCLUSION: The present study revealed that supplementation of tris basic extender with trehalose at 100 mM and or IGF-1 at 100 ng/mL or 150 ng per mL improves the post-thaw semen characteristics and protects certain fertility related sperm membrane proteins. Doi.org/10.54680/fr23510110612.

[Effect of recombinant human growth hormone on serum Klotho and fibroblast growth factor 23 in children with idiopathic short stature]. / é‡ç»„äººç”Ÿé•¿æ¿€ç´ æ²»ç–—å¯¹ç‰¹å‘æ€§çŸ®å°ç—‡å„¿ç«¥è¡€æ¸ Klothoå’Œæˆçº¤ç»´ç»†èƒžç”Ÿé•¿å› å­23的影响.

OBJECTIVES: To investigate the changes in the serum levels of Klotho, fibroblast growth factor 23 (FGF23), and insulin-like growth factor-1 (IGF-1) in children with idiopathic short stature (ISS) before and after recombinant human growth hormone (rhGH) treatment, as well as the correlation of Klotho and FGF23 with the growth hormone (GH)/IGF-1 growth axis in these children. METHODS: A prospective study was conducted on 33 children who were diagnosed with ISS in the Department of Pediatrics, Hebei Provincial People's Hospital, from March 10, 2021 to December 1, 2022 (ISS group). Twenty-nine healthy children, matched for age and sex, who attended the Department of Child Healthcare during the same period, were enrolled as the healthy control group. The children in the ISS group were treated with rhGH, and the serum levels of Klotho, FGF23, and IGF-1 were measured before treatment and after 3, 6, and 9 months of treatment. A correlation analysis was conducted on these indexes. RESULTS: There were no significant differences in the serum levels of IGF-1, Klotho, and FGF23 between the ISS and healthy control groups (P>0.05). The serum levels of Klotho, FGF23, and IGF-1 increased significantly in the ISS group after 3, 6, and 9 months of rhGH treatment (P<0.05). In the ISS group, Klotho and FGF23 levels were positively correlated with the phosphate level before treatment (P<0.05). Before treatment and after 3, 6, and 9 months of rhGH treatment, the Klotho level was positively correlated with the IGF-1 level (P<0.05), the FGF23 level was positively correlated with the IGF-1 level (P<0.05), and the Klotho level was positively correlated with the FGF23 level (P<0.05), while Klotho and FGF23 levels were not correlated with the height standard deviation of point (P>0.05). CONCLUSIONS: The rhGH treatment can upregulate the levels of Klotho, FGF23, and IGF-1 and realize the catch-up growth in children with ISS. Klotho and FGF23 may not directly promote the linear growth of children with ISS, but may have indirect effects through the pathways such as IGF-1 and phosphate metabolism. The consistent changes in Klotho, FGF23 and IGF-1 levels show that there is a synergistic relationship among them in regulating the linear growth of ISS children.