ATI-2341 Trifluoroacetic Acid (TFA) Promotes Menstrual Blood-Derived Mesenchymal Stem Cell Recruitment and Enhances Uterine Repair Through Gi Pathway in Asherman’s Syndrome

This study aimed to explore the role of ATI-2341 in Asherman’s syndrome and its impact on menstrual blood-derived mesenchymal stem cells (MenSCs). Following establishment of endometrial injury model, MenSCs were extracted from rats and cultured. They were treated with ATI-2341 TFA at different concentrations (10 ng/mL, 50 ng/mL, 100 ng/mL) and MenSCs treated without ATI-2341 TFA were taken as controls. Flow cytometry was conducted to detect the cell cycle. MTT was carried out to evaluate proliferation of endometrial cells. The expression levels of MMP-9, TIMP-1, CK, and VIM were determined with staining used to reflect morphology of endometrium. Administration with ATI-2341 TFA resulted in decreased expression of MMP-9 and increased expression of TIMP-1 in a dose-dependent manner. Of note, the increase of ATI-2341 TFA concentration was accompanied with elevated cell proliferation rate, increased number of glands in the endometrium, and decreased fibrosis area. As treated with 100 ng/mL ATI-2341 TFA, the cells exhibited more glands than that under other concentrations with uniformly arranged glands and lowest expression levels of CK and VIM, control group had plenty of blue-stained collagen fibers in the intima and least amount of glands. ATI-2341 TFA 100 ng/mL induced endometrial epithelial recruitment effect on MenSCs and promoted endometrial repair more significantly than Gi-3 pathway agonists. Collectively, ATI-2341 TFA enhances MenSC recruitment and facilitates endometrial epithelial cells proliferation and the repair of uterine damage in Asherman’s syndrome through Gi pathway. These findings provide a\ novel insight into the MenSC-based treatment against Asherman’s syndrome and deserve further investigation.

A novel mouse model of type 2N VWD was developed by CRISPR/Cas9 gene editing and recapitulates human type 2N VWD

Type 2N von Willebrand disease is caused by mutations in the factor VIII (FVIII) binding site of von Willibrand factor (VWF), resulting in dysfunctional VWF with defective binding capacity for FVIII. Here we developed a novel type 2N mouse model using CRISPR/Cas9 technology. In homozygous VWF2N/2N mice, plasma VWF levels were normal (1167±257 mU/ml) but the VWF was completely incapable of binding FVIII, resulting in 53±23 mU/ml of plasma FVIII levels that were similar to those in VWF deficient (VWF-/-) mice. When wild-type human or mouse VWF was infused into VWF2N/2N mice, endogenous plasma FVIII was restored, peaking at 4-6 hours post-infusion, demonstrating that FVIII expressed in VWF2N mice is viable, but short-lived unprotected in plasma due to dysfunctional 2N-VWF. The whole blood clotting time and thrombin generation were impaired in VWF2N/2N but not in VWF-/- mice. The bleeding time and blood loss in VWF2N/2N mice were similar to wild-type mice in the lateral tail vein or ventral artery injury model. However, VWF2N/2N, but not VWF-/- mice, lost a significant amount of blood during the primary bleeding phase after a tail tip amputation injury model, indicating that there are other alternative pathway(s) that can at least partially restore hemostasis when VWF is absent. In summary, we have developed a novel mouse model by gene editing with both the pathophysiology and clinical phenotype found in severe type 2N patients. This unique model can be used to investigate the biological properties of VWF/FVIII association in hemostasis and beyond.

Nonwoven-based gelatin/polycaprolactone membrane loaded with ERK inhibitor U0126 for treatment of tendon defects

Background Tendon is a major component of musculoskeletal system connecting the muscles to the bone. Tendon injuries are very common orthopedics problems leading to impeded motion. Up to now, there still lacks effective treatments for tendon diseases. Methods Tendon stem/progenitor cells (TSPCs) were isolated from the patellar tendons of SD rats. The expression levels of genes were evaluated by quantitative RT-PCR. Immunohistochemistry staining was performed to confirm the presence of tendon markers in tendon tissues. Bioinformatics analysis of data acquired by RNA-seq was used to find out the differentially expressed genes. Rat patellar tendon injury model was used to evaluate the effect of U0126 on tendon injury healing. Biomechanical testing was applied to evaluate the mechanical properties of newly formed tendon tissues. Results In this study, we have shown that ERK inhibitor U0126 rather PD98059 could effectively increase the expression of tendon-related genes and promote the tenogenesis of TSPCs in vitro. To explore the underlying mechanisms, RNA sequencing was performed to identify the molecular difference between U0126-treated and control TSPCs. The result showed that GDF6 was significantly increased by U0126, which is an important factor of the TGFβ superfamily regulating tendon development and tenogenesis. In addition, NBM (nonwoven-based gelatin/polycaprolactone membrane) which mimics the native microenvironment of the tendon tissue was used as an acellular scaffold to carry U0126. The results demonstrated that when NBM was used in combination with U0126, tendon healing was significantly promoted with better histological staining outcomes and mechanical properties. Conclusion Taken together, we have found U0126 promoted tenogenesis in TSPCs through activating GDF6, and NBM loaded with U0126 significantly promoted tendon defect healing, which provides a new treatment for tendon injury.

Hypocrates is a genetically encoded fluorescent biosensor for (pseudo)hypohalous acids and their derivatives

The lack of tools to monitor the dynamics of (pseudo)hypohalous acids in live cells and tissues hinders a better understanding of inflammatory processes. Here we present a fluorescent genetically encoded biosensor, Hypocrates, for the visualization of (pseudo)hypohalous acids and their derivatives. Hypocrates consists of a circularly permuted yellow fluorescent protein integrated into the structure of the transcription repressor NemR from Escherichia coli. We show that Hypocrates is ratiometric, reversible, and responds to its analytes in the 106 M−1s−1 range. Solving the Hypocrates X-ray structure provided insights into its sensing mechanism, allowing determination of the spatial organization in this circularly permuted fluorescent protein-based redox probe. We exemplify its applicability by imaging hypohalous stress in bacteria phagocytosed by primary neutrophils. Finally, we demonstrate that Hypocrates can be utilized in combination with HyPerRed for the simultaneous visualization of (pseudo)hypohalous acids and hydrogen peroxide dynamics in a zebrafish tail fin injury model.

Butylphthalide Inhibits TLR4/NF-κB Pathway by Upregulation of miR-21 to Have the Neuroprotective Effect

Stroke is a disease with the highest incidence rate and the highest mortality rate in the world. The study aims to verify the neuroprotective effect of Butylphthalide. The mice were divided into sham group, MCAO group, and MCAO + Butylphthalide-treated group. The mice in MCAO + Butylphthalide-treated group were administered with 70 mg/kg Butylphthalide injection intraperitoneally after cerebral ischemia-reperfusion. The normal saline with the same volume was administered intraperitoneally for the mice in the MCAO group and sham group. The levels of miR-21 in brain tissue and cells were detected by qPCR. The OGD/R injury model of Neuro2A cells was used to simulate the hypoxic-ischemic environment of neurons in vitro. The proliferation rate of Neuro2A cells was detected with CCK-8. The production of ROS was detected with DCFH-DA. Compared with the mice in MCAO group, a decrease ( P < 0.01 ) was observed in the functional neurologic impairment scoring, cerebral infarction volume, and brain loss volume in the mice treated with MCAO + Butylphthalide, but an increase ( P < 0.01 ) was observed in the level of miR-21, which was positively correlated with functional neurologic impairment scoring (r = −0.8933, P < 0.001 ). MTT assay showed that the cell viability of OGD/R + Butylphthalide group was significantly higher than that of other groups ( P < 0.001 ), and the activity of ROS was significantly decreased ( P < 0.001 ). The WB results showed that, compared with OGD/R + miR-NC and control groups, the ratio of Bcl-2/Bax in OGD/R + Butylphthalide group and OGD/R + miR-21 mimics group was significantly higher ( P < 0.05 ), while the ratio of caspase-3/GAPDH was significantly lower ( P < 0.05 ). In conclusion, Butylphthalide has neuroprotective effect on the mouse model of MCAO. It may upregulate the level of miR-21 to inhibit neuronal apoptosis and ROS production and improve the proliferation activity. The specific mechanism may lie in inhibiting TLR4/NF-κB pathway.

Computer Intelligent Algorithm in the Recovery of the Elbow Joint Sports Injury Model

In this study, the inverse kinematics mathematics computer intelligent algorithm model is used to study the sports injuries of the elbow joint of adolescents. At the same time, we simulated the movement parameter changes during the rehabilitation training of the patient’s wrist and proposed a joint angular velocity function based on cubic fitting. Research has found that when the training scene changes greatly or the target task is changed, the smoothness of the elbow joint movement will change. The research conclusions of this article provide a theoretical basis for the selection of man-machine action points and the formulation of rehabilitation training methods. This article establishes the degree-of-freedom simulation model of the operating arm, which is the number of independent position variables in the operating arm, and these position variables determine the positions of all parts in the mechanism.

Citrus Naringenin Increases Neuron Survival in Optic Nerve Crush Injury Model by Inhibiting JNK-JUN Pathway

Traumatic nerve injury activates cell stress pathways, resulting in neuronal death and loss of vital neural functions. To date, there are no available neuroprotectants for the treatment of traumatic neural injuries. Here, we studied three important flavanones of citrus components, in vitro and in vivo, to reveal their roles in inhibiting the JNK (c-Jun N-terminal kinase)-JUN pathway and their neuroprotective effects in the optic nerve crush injury model, a kind of traumatic nerve injury in the central nervous system. Results showed that both neural injury in vivo and cell stress in vitro activated the JNK-JUN pathway and increased JUN phosphorylation. We also demonstrated that naringenin treatment completely inhibited stress-induced JUN phosphorylation in cultured cells, whereas nobiletin and hesperidin only partially inhibited JUN phosphorylation. Neuroprotection studies in optic nerve crush injury mouse models revealed that naringenin treatment increased the survival of retinal ganglion cells after traumatic optic nerve injury, while the other two components had no neuroprotective effect. The neuroprotection effect of naringenin was due to the inhibition of JUN phosphorylation in crush-injured retinal ganglion cells. Therefore, the citrus component naringenin provides neuroprotection through the inhibition of the JNK-JUN pathway by inhibiting JUN phosphorylation, indicating the potential application of citrus chemical components in the clinical therapy of traumatic optic nerve injuries.

Hepatoprotective Effect of Cereal Vinegar Sediment in Acute Liver Injury Mice and Its Influence on Gut Microbiota

Cereal vinegar sediment (CVS) is a natural precipitate formed during the aging process of traditional grain vinegar. It has been used as Chinese traditional medicine, while its composition and function are reported minimally. In this study, we measured CVS in terms of saccharide, protein, fat and water content, and polyphenol and flavonoid content. Furthermore, we determined the amino acids, organic acids, and other soluble metabolites in CVS using reverse-phase high-performance liquid chromatography (RP-HPLC), HPLC, and liquid chromatography with tandem mass spectrometry (LC-MS/MS) platforms. The hepatoprotective effect of CVS was evaluated in acute CCl4-induced liver injury mice. Administration of CVS for 7 days prior to the CCl4 treatment can significantly decrease liver alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels and reactive oxygen species (ROS) levels, compared with those in the hepatic injury model group. The gut microbiota was changed by CCl4 administration and was partly shifted by the pretreatment of CVS, particularly the Muribaculaceae family, which was increased in CVS-treated groups compared with that in the CCl4 administration group. Moreover, the abundances of Alistipes genus and Muribaculaceae family were correlated with the liver ALT, AST, and malondialdehyde (MDA) levels. Our results illustrated the composition of CVS and its hepatoprotective effect in mice, suggested that CVS could be developed as functional food to prevent acute liver injury.

Neonatal enthesis healing involves non-inflammatory formation of acellular scar through ECM secretion by resident cells

Wound healing is a well-orchestrated process that typically recruits the immune and vascular systems to restore the structure and function of the injured tissue. Injuries to the enthesis, a hypocellular and avascular tissue, often result in fibrotic scar formation and loss of mechanical properties, thereby severely affecting musculoskeletal function and life quality. This raises questions about the healing capabilities of the enthesis. Here, we established an injury model to the Achilles entheses of neonatal mice to study the possibility that at an early age, the enthesis can heal more effectively. Histology and immunohistochemistry analyses revealed an atypical process that did not involve inflammation or angiogenesis. Instead, neonatal enthesis healing was mediated by secretion of collagen types I and II by resident cells, which formed a permanent hypocellular and avascular scar. Transmission electron microscopy showed that the cellular response to injury, including ER stress, autophagy and cell death, varied between the tendon and cartilage ends of the enthesis. Single-molecule in situ hybridization, immunostaining, and TUNEL assays verified these differences. Finally, gait analysis showed that these processes effectively restored function of the injured leg. Collectively, these findings reveal a novel healing mechanism in neonatal entheses, whereby local ECM secretion by resident cells forms an acellular ECM deposit in the absence of inflammation markers, allowing gait restoration. These insights into the healing mechanism of a complex transitional tissue may lead to new therapeutic strategies for adult enthesis injuries.

PuraStat RADA16 Self-Assembling Peptide Reduces Postoperative Abdominal Adhesion Formation in a Rabbit Cecal Sidewall Injury Model

Objective: To evaluate the effect of PuraStat (2.5% RADA16) administration on postoperative abdominal adhesion formation in an in vivo model.Methods: Anesthetized New Zealand white rabbits underwent cecal sidewall abrasion surgery in which the cecal serosa and juxtaposed parietal peritoneum were abraded after access through an abdominal midline incision. Eight animals were randomized to receive PuraStat administration at the interface of the injured tissues before incision closure, and five animals served as untreated controls. Treated animals received 3–12 ml PuraStat solution per lesion. Animals were sacrificed 14 days after surgery and examined for adhesion formation at the wound site.Results: At study terminus, adhesions were identified in 90% (9/10) of abraded cecum/peritoneal wound sites in untreated controls versus 25% (4/16) of PuraStat-treated sites (p = 0.004). Mean ± SD Total Adhesion Score (average of the values for extent + strength of the adhesion in both defects per animal; maximum score = 14 points) was significantly 76% lower in PuraStat-treated animals (2.0 ± 3.0 points) compared to untreated controls (8.2 ± 1.9 points) (p = 0.029). Mean adhesion coverage area of wound sites was 79% lower in PuraStat-treated animals than controls (p &lt; 0.001), and mean adhesion durability was 72% lower in PuraStat-treated animals versus controls (p = 0.005). Remnant hydrogel was observed at the wound sites of 75% of treated animals at postoperative Day 14.Conclusion: PuraStat treatment has a positive protective effect in the cecal sidewall injury model, and significantly reduces abdominal adhesion formation at the interface of the injured cecum and overlying peritoneal sidewall defect.