Establishment and comparison of different procedures for modeling intrauterine adhesion in rats: A preliminary study.

The establishment of a stable animal model for intrauterine adhesion (IUA) can significantly enhance research on the pathogenesis and pathological changes of this disease, as well as on the development of innovative therapeutic approaches. In this study, three different modeling methods, including phenol mucilage combined mechanical scraping, ethanol combined mechanical scraping and ethanol modeling alone were designed. The morphological characteristics of the models were evaluated. The underlying mechanisms and fertility capacity of the ethanol modeling group were analyzed and compared to those of the sham surgery group. All three methods resulted in severe intrauterine adhesions, with ethanol being identified as a reliable modeling agent and was subsequently subjected to further evaluation. Immunohistochemistry and RT-PCR results indicated that the ethanol modeling group exhibited an increase in the degree of fibrosis and inflammation, as well as a significant reduction in endometrial thickness, gland number, vascularization, and endometrial receptivity, ultimately resulting in the loss of fertility capacity. The aforementioned findings indicate that the intrauterine perfusion of 95 % ethanol is efficacious in inducing the development of intrauterine adhesions in rats. Given its cost-effectiveness, efficacy, and stability in IUA formation, the use of 95 % ethanol intrauterine perfusion may serve as a novel platform for evaluating innovative anti-adhesion materials and bioengineered therapies.

Bioactive ECM-Loaded SIS Generated by the Optimized Decellularization Process Exhibits Skin Wound Healing Ability in Type I Diabetic Rats.

Patients with diabetes have 15-25% chance for developing diabetic ulcers as a severe complication and formidable challenge for clinicians. Conventional treatment for diabetic ulcers is to surgically remove the necrotic skin, clean the wound, and cover it with skin flaps. However, skin flap often has a limited efficacy, and its acquisition requires a second surgery, which may bring additional risk for the patient. Skin tissue engineering has brought a new solution for diabetic ulcers. Herein, we have developed a bioactive patch through a compound culture and the optimized decellularization strategy. The patch was prepared from porcine small intestinal submucosa (SIS) and modified by an extracellular matrix (ECM) derived from urine-derived stem cells (USCs), which have low immunogenicity while retaining cytokines for angiogenesis and tissue regeneration. The protocol included the optimization of the decellularization time and the establishment of the methods. Furthermore, the in vitro mechanism of wound healing ability of the patch was investigated, and its feasibility for skin wound healing was assessed through an antishrinkage full-thickness skin defect model in type I diabetic rats. As shown, the patch displayed comparable effectiveness to the USCs-loaded SIS. Our findings suggested that this optimized decellularization protocol may provide a strategy for cell-loaded scaffolds that require the removal of cellular material while retaining sufficient bioactive components in the ECM for further applications.

Application of antibody-conjugated small intestine submucosa to capture urine-derived stem cells for bladder repair in a rabbit model.

The need for bladder reconstruction and side effects of cystoplasty have spawned the demand for the development of alternative material substitutes. Biomaterials such as submucosa of small intestine (SIS) have been widely used as patches for bladder repair, but the outcomes are not fully satisfactory. To capture stem cells in situ has been considered as a promising strategy to speed up the process of re-cellularization and functionalization. In this study, we have developed an anti-CD29 antibody-conjugated SIS scaffold (AC-SIS) which is capable of specifically capturing urine-derived stem cells (USCs) in situ for tissue repair and regeneration. The scaffold has exhibited effective capture capacity and sound biocompatibility. In vivo experiment proved that the AC-SIS scaffold could promote rapid endothelium healing and smooth muscle regeneration. The endogenous stem cell capturing scaffolds has thereby provided a new revenue for developing effective and safer bladder patches.

Procyanidins-crosslinked small intestine submucosa: A bladder patch promotes smooth muscle regeneration and bladder function restoration in a rabbit model.

Currently the standard surgical treatment for bladder defects is augmentation cystoplasty with autologous tissues, which has many side effects. Biomaterials such as small intestine submucosa (SIS) can provide an alternative scaffold for the repair as bladder patches. Previous studies have shown that SIS could enhance the capacity and compliance of the bladder, but its application is hindered by issues like limited smooth muscle regeneration and stone formation since the fast degradation and poor mechanical properties of the SIS. Procyanidins (PC), a natural bio-crosslinking agent, has shown anti-calcification, anti-inflammatory and anti-oxidation properties. More importantly, PC and SIS can crosslink through hydrogen bonds, which may endow the material with enhanced mechanical property and stabilized functionalities. In this study, various concentrations of PC-crosslinked SIS (PC-SIS) were prepared to repair the full-thickness bladder defects, with an aim to reduce complications and enhance bladder functions. In vitro assays showed that the crosslinking has conferred the biomaterial with superior mechanical property and anti-calcification property, ability to promote smooth muscle cell adhesion and upregulate functional genes expression. Using a rabbit model with bladder defects, we demonstrated that the PC-SIS scaffold can rapidly promote in situ tissue regrowth and regeneration, in particular smooth muscle remodeling and improvement of urinary functions. The PC-SIS scaffold has therefore provided a promising material for the reconstruction of a functional bladder.

Hypoxic preconditioning of human urine-derived stem cell-laden small intestinal submucosa enhances wound healing potential.

BACKGROUND: Urine-derived stem cells (USCs) are a valuable stem cell source for tissue engineering because they can be harvested non-invasively. Small intestine submucosa (SIS) has been used as scaffolds for soft tissue repair in the clinic. However, the feasibility and efficacy of a combination of USCs and SIS for skin wound healing has not been reported. In this study, we created a tissue-engineered skin graft, termed the SIS+USC composite, and hypothesized that hypoxic preconditioning would improve its wound healing potential. METHODS: USCs were seeded on SIS membranes to fabricate the SIS+USC composites, which were then cultured in normoxia (21% O2) or preconditioned in hypoxia (1% O2) for 24 h, respectively. The viability and morphology of USCs, the expression of genes related to wound angiogenesis and reepithelialization, and the secretion of growth factors were determined in vitro. The wound healing ability of the SIS+USC composites was evaluated in a mouse full-thickness skin wound model. RESULTS: USCs showed good cell viability and morphology in both normoxia and hypoxic preconditioning groups. In vitro, hypoxic preconditioning enhanced not only the expression of genes related to wound angiogenesis (VEGF and Ang-2) and reepithelialization (bFGF and EGF) but also the secretion of growth factors (VEGF, EGF, and bFGF). In vivo, hypoxic preconditioning significantly improved the wound healing potential of the SIS+USC composites. It enhanced wound angiogenesis at the early stage of wound healing, promoted reepithelialization, and improved the deposition and remodeling of collagen fibers at the late stage of wound healing. CONCLUSIONS: Taken together, this study shows that hypoxic preconditioning provides an easy and efficient strategy to enhance the wound healing potential of the SIS+USC composite.

Copper promotes the migration of bone marrow mesenchymal stem cells via Rnd3-dependent cytoskeleton remodeling.

The motility of mesenchymal stem cells (MSCs) is highly related to their homing in vivo, a critical issue in regenerative medicine. Our previous study indicated copper (Cu) might promote the recruitment of endogenous MSCs in canine esophagus defect model. In this study, we investigated the effect of Cu on the motility of bone marrow mesenchymal stem cells (BMSCs) and the underlying mechanism in vitro. Cu supplementation could enhance the motility of BMSCs, and upregulate the expression of hypoxia-inducible factor 1α (Hif1α) at the protein level, and upregulate the expression of rho family GTPase 3 (Rnd3) at messenger RNA and protein level. When Hif1α was silenced by small interfering RNA (siRNA), Cu-induced Rnd3 upregulation was blocked. When Rnd3 was silenced by siRNA, the motility of BMSCs was decreased with or without Cu supplementation, and Cu-induced cytoskeleton remodeling was neutralized. Furthermore, overexpression of Rnd3 also increased the motility of BMSCs and induced cytoskeleton remodeling. Overall, our results demonstrated that Cu enhanced BMSCs migration through, at least in part, cytoskeleton remodeling via Hif1α-dependent upregulation of Rnd3. This study provided an insight into the mechanism of the effect of Cu on the motility of BMSCs, and a theoretical foundation of applying Cu to improve the recruitment of BMSCs in tissue engineering and cytotherapy.

Identification and characterization of two morphologically distinct stem cell subpopulations from human urine samples.

Urine-derived stem cells (USCs) have shown potentials for the treatment of skeletal and urological disorders. Based on published literature and our own data, USCs consist of heterogeneous populations of cells. In this paper, we identify and characterize two morphologically distinct subpopulations of USCs from human urine samples, named as spindle-shaped USCs (SS-USCs) and rice-shaped USCs (RS-USCs) respectively. The two subpopulations showed similar clone-forming efficiency, while SS-USCs featured faster proliferation, higher motility, and greater potential for osteogenic and adipogenic differentiation, RS-USCs showed greater potential for chondrogenic differentiation. POU5F1 was strongly expressed in both subpopulations, but MYC was weakly expressed. Both subpopulations showed similar patterns of CD24, CD29, CD34, CD44, CD73, CD90 and CD105 expression, while a higher percentage of RS-USCs were positive for CD133. SS-USCs were positive for VIM, weakly positive for SLC12A1 and UMOD, and negative for KRT18, NPHS1, AQP1 and AQP2, indicating a renal mesenchyme origin; while RS-USCs are positive for VIM, partially positive for KRT18, NPHS1, AQP1, SLC12A1 and UMOD, and negative for AQP2, indicating a nephron tubule origin. The above results can facilitate understanding of the biological characteristics of subpopulations of USCs, and provide a basis for further research and applications of such cells.

Collagen Hydrogel Functionalized with Collagen-Targeting IFNA2b Shows Apoptotic Activity in Nude Mice with Xenografted Tumors.

Interferon alpha 2b (IFNA2b) has been used in immunotherapy for cancers with certain success. To reduce fast diffusion of IFNA2b and consequent dose-dependent side effects, we constructed a collagen hydrogel loaded with IFNA2b fused to collagen-binding domain by using methods of tissue engineering. The fusion protein showed apoptotic activity similar to that of native IFNA2b against MCF-7 cells in vitro, but with relatively higher affinity for collagen type I. Accordingly, the former diffused out of the collagen matrix slower than the latter. Importantly, collagen hydrogels loaded with the fusion protein possessed apoptotic activity in vitro and released the engineered cytokine in a controlled manner. In addition, such hydrogels reduced tumor size and extended the survival of the mouse model with xenografted tumors, which suggested a moderate antitumor activity in vivo.

Epigallocatechin-3-gallate Cross-Linked Small Intestinal Submucosa for Guided Bone Regeneration.

Collagen membranes are widely used in guided bone regeneration (GBR) because of their good biocompatibility and low immunogenicity. As a bioderived collagen membrane, small intestinal submucosa (SIS) has good regenerative potential for soft tissue repair, but it lacks sufficient mechanical properties for GBR application unless properly modifided. Epigallocatechin-3-gallate (EGCG) is a natural cross-linking agent featuring osteoinductive activity. In this study, we modified SIS by EGCG cross-linking, and such modified materials were characterized both in vitro and in vivo. The results showed that EGCG cross-linking significantly improved the mechanical properties and hydrophilicity of SIS while maintaing good cytocompatibility. Compared to SIS, EGCG-cross-linked SIS (E-SIS) enhanced the adhesion of fibroblasts and preosteoblasts and promoted the osteogenic differentiation of MC3T3-E1 cells cultured on the materials. In a rat cranial defect model, E-SIS material showed better occlusion effect than SIS material. Most importantly, E-SIS material accelerated bone regeneration more than SIS material and even a commercially available GBR membrane. Taken together, we conclude that E-SIS is a promising material as a GBR membrane.

Comparison of small intestinal submucosa and polypropylene mesh for abdominal wall defect repair.

Abdominal wall defects are a common medical problem, and inadequate repair methods can lead to serious complications. Abdominal wall reconstruction using autologous tissue, or non-biological, biological, or composite patches is often performed to repair defective areas. In particular, composite patches containing both polymeric and biological materials have gained increasing attention due to their good mechanical properties and biocompatibility. However, it is still unclear whether the quality of repairs using composite patches is superior to that of a biological patch. Based on the limitations of previous studies, we compared small intestinal submucosa (SIS) patches with SIS + polypropylene mesh (PPM) patches for repairing abdominal wall defects in adult beagle dogs. Forty-five female dogs were subjected to surgical resection to produce abdominal wall defects. SIS or SIS + PPM was used as patch for the defects. Morphology, biomechanics, and histological evaluations were performed to evaluate the efficacy and safety of such therapies. Our findings demonstrated that SIS had advantages over SIS + PPM considering biological activity and histocompatibility without increasing the risk of repair failure.

Altered gene expression profile in a rat model of gentamicin-induced ototoxicity and nephrotoxicity, and the potential role of upregulated Ifi44 expression.

As demonstrated by Alport syndrome, the co­occurrence of auditory and urinary system malformations, and gentamicin-induced ototoxicity and nephrotoxicity, the ears and kidneys potentially share certain molecular pathways. In the present study, microarray chips were used to analyze the changes in the gene expression profile using a rat model of gentamicin­induced ototoxicity and nephrotoxicity, using rat liver tissue as a control. A number of genes were identified to exhibit similar expression changes in the rat ears and kidney tissues, among which microtubule­associated protein 44 (Ifi44), was selected for further analysis to validate its expression changes and confirm potential involvement in the inflammation process in the disease model. Ifi44 is a member of the type I interferon­inducible gene family. Reverse transcription­quantitative polymerase chain reaction, western blotting and immunohistochemistry were performed; the results demonstrated that more inflammatory cells were present in cochlear and renal parenchyma in gentamycin­induced rats, and Ifi44 expression was increased in these two organs compared with control rats. Taken together, with its role in lupus nephritis and expression in the inner ear, the results suggested that Ifi44 is potentially involved in the inflammation associated with gentamicin­induced ototoxicity and nephrotoxicity. The approach of the current study has also provided a strategy for delineating common pathways shared by organs involved in specific diseases.