Small Intestinal Submucosa Matrix Research Articles

Healing potential of chitosan and decellularized intestinal matrix with mesenchymal stem cells and growth factor in burn wound in rat

Abstract Background Burns are one of the most common and devastating forms of trauma. Patients with serious thermal injury require immediate specialized care in order to minimize morbidity and mortality. The objective of this study was to evaluate the healing potential of porcine decellularized small intestinal submucosa (SIS) matrix and chitosan with mesenchymal stem cells (rMSC) and epidermal growth factor (mEGF) on the healing of full thickness burn wound in rat model. Methods One 2 × 2 cm2 size full thickness burn wound was created on the dorsum of each animal under general anaesthesia. Burn wounds were treated with silver sulfadiazine; medical grade chitosan; chitosan with rMSC; chitosan with rMSC and mEGF; decellularized porcine SIS; rMSC seeded with SIS; rMSC seeded with SIS and mEGF in groups A, B, C, D, E, F and G, respectively. Healing was evaluated on the basis of percentage of wound contraction, colour digital imaging, immunological, histopathological, immunohistochemistry and gelatin zymography observations. Results A decrease in wound area was noticed in stem cell and m-EGF treated groups. Early angiogenesis and epithelialisation was observed in groups, where decellularized SIS matrix was combined with r-MSC and m-EGF. Type III collagen was found more in control group by immunohistochemistry which indicated abnormal collagen deposition. Gelatin zymography revealed expression of MMPs in the control group suggesting a chronic nature of wound. Chitosan in combination of rMSC and mEGF proved to be efficient for full-thickness burn wound healing. Conclusion Decellularized SIS matrix when combined with r-MSC and m-EGF has healing potential and showed best healing response than conventional treatment for the reconstruction of full-thickness skin burn wounds in rats.

The cytoptrotection of small intestinal submucosa-derived gel in HL-1 cells during hypoxia/reoxygenation-induced injury.

Small intestinal submucosa (SIS)-derived gel injected into infarcted myocardium has been shown to promote repair and regeneration after myocardial infarction (MI); however, the specific impact of SIS gel on cardiomyocytes remained unknown. The aim of this study was to characterise SIS gel function in hypoxia-reoxygenation (H/R)-induced cardiomyocyte damage and its potential mechanism. HL-1 cardiomyocytes seeded on SIS matrix-coated plates, SIS gel, and uncoated plates were subjected to H/R, cell viability, apoptosis, expression of caspase-3, Bcl-2, and Bax were investigated. SIS gel and SIS matrix as coating substrates markedly improved cell viability, preventing cell apoptosis compared with uncoated plates, with SIS gel yielding the best cytoprotective effects. SIS gel down-regulated expression of pro-inflammatory cytokines (TNF-α, CCL2, and IL-6) by inhibiting the JNK-mitogen-activated protein kinase (MAPK)/NF-κB pathways. Furthermore, SIS gel protected cardiomyocytes from apoptosis by activating protein kinase B (AKT) and extracellular-signal-regulated kinase (ERK) pathways, and markedly up-regulated antiapoptotic Bcl-2 expression but inhibited that of proapoptotic Bax and c-caspase 3. Together, these findings show that SIS gel could decrease H/R-induced cell apoptosis through a mechanism potentially related to its ability to regulate expression of inflammatory cytokines and antiapoptosis signalling pathways to prevent cell apoptosis. Our findings thereby shed light on the mechanism related to SIS gel therapeutic efficacy for MI.

Ureteral reconstruction with decellularized small intestinal submucosa matrix for ureteral stricture: A preliminary report of two cases

ObjectiveTo determine the feasibility of decellularized small intestinal submucosa (SIS) matrix in repairing ureteral strictures. MethodsTwo patients with ureteral stenoses underwent ureteral reconstruction with SIS matrix at the Zhejiang Provincial Corps Hospital of Chinese People's Armed Forces between June 2014 and June 2016. The ureteral stenoses were repaired with a semi-tubular SIS matrix and the postoperative recoveries were observed. ResultsBoth operations were successfully completed. The average operative time was 90 min and the average length of hospital stay was 15 days. No fevers, incision infections, intestinal obstruction, graft rejection, or other serious complications were noted. After 2 months, ureteroscopic examinations showed that the surfaces of the original patches were covered by mucosa and there were no apparent stenoses in the lumens. The ureteral stents were replaced every 2 months postoperatively and removed 12 months postoperatively. No infections or urinary leakage occurred after removal of the stents. Intravenous urography was performed 6 and 12 months postoperatively. The results showed that the ureters were not obstructed and there was no apparent stenosis at the anastomosis sites. The average follow-up time was >12 months. Long-term follow-up is still ongoing, and computed tomography examinations of the urinary tract have been conducted in the outpatient department of our hospital 1, 3, and 6 months after removal of the double-J stents, suggesting the absence of hydronephrosis. The serum creatinine levels remained stable during the follow-up. ConclusionSIS matrix reconstruction is a feasible method to repair ureters stenosis.

Small Intestinal Submucosa Matrix as a Novel Therapy for Wounds in Dystrophic Epidermolysis Bullosa

Small Intestinal Submucosa Matrix as a Novel Therapy for Wounds in Dystrophic Epidermolysis Bullosa

Minimal modulation of the host immune response to SIS matrix implants by mesenchymal stem cells from the amniotic fluid

Surgical restoration of soft tissue defects often requires implantable devices. The clinical outcome of the surgery is determined by the properties inherent to the used matrix. Mesenchymal stem cells (MSC) modulate the immune processes after in vivo transplantation and their addition to matrices is associated with constructive remodeling. Herein we evaluate the potential of MSC derived from the amniotic fluid(AF-MSC), an interesting MSC source for cell therapeutic applications in the perinatal period, for immune modulation when added to a biomaterial. We implant cell free small intestinal submucosa (SIS) or SIS seeded with AF-MSC at a density of 1×105/cm2 subcutaneously at the abdominal wall in immune competent rats. The host immune response is evaluated at 3, 7 and 14days postoperatively. The matrix-specific or cellular characteristics are not altered after 24h of in vitro co-culture of SIS with AF-MSC. The host immune response was not different between animals implanted with cell free or AF-MSC-seeded SIS in terms of cellular infiltration, vascularity, macrophage polarization or scaffold replacement. Profiling the mRNA expression level of inflammatory cytokines at the matrix interface shows a significant reduction in the expression of the pro-inflammatory marker Tnf-α and a trend towards lower iNos expression upon AF-MSC-seeding of the SIS matrix. Anti-inflammatory marker expression does not alter upon cell seeding of matrix implants. We conclude that SIS is a suitable substrate for in vitro culture of AF-MSC and fibroblasts. AF-MSC addition to SIS does not significantly modulate the host immune response after subcutaneous implantation in rats.

Preparation and characterization of pro-angiogenic gel derived from small intestinal submucosa

Gels derived from decellularized small intestinal submucosa (SIS) have been used to repair ischemic myocardium and deliver protein drug. However, their material properties and effects on cell behavior are not well understood, in part because of the difficulty of gelling in vitro. In this study, soluble SIS matrix, which was easily handled and could effectively gel, was successfully prepared using a modified method. Fourier transform infrared spectroscopy confirmed that the SIS gel contained not only collagen but also sulfated glycosaminoglycans (sGAGs). Interestingly, the sustained release of vascular endothelial growth factor and basic fibroblast growth factor within the SIS gel was detected, and no initial burst release was observed. The SIS gel was more capable of evoking neovascularization than collagen type I gel, as determined by tube formation experiments in human umbilical vein endothelial cells, the mouse aortic ring assay, and animal experiments. The upregulated expression of kinase insert domain receptor (KDR), Notch1, and Ang2, the key genes in angiogenesis that were evaluated in HUVECs seeded on the SIS gel, confirmed that angiogenesis bioactive factors contained in the SIS gel are indeed active and effective. The SIS gel significantly promoted neovascularization compared to the collagen type I gel in vivo. Histology revealed adequate host tissue response in engraftment both types of gels. Together, these data demonstrate that the SIS gel is a promising and attractive candidate for tissue engineering, especially in promoting vessel formation. Statement of significanceThe material properties of small intestinal submucosa (SIS) gel and the effect of these properties upon cell behavior are not well understood, in part due to the difficulty of gelling in vitro. In this study, soluble SIS matrix, which was easily handled and gelled was prepared using modified method. The material properties and biocompatibility of SIS gel were explored. The sustained release of growth factors from this gel was observed along with its degradation in vitro. The results demonstrate that the SIS gel promote angiogenesis in vitro and in vivo. The SIS gel biological properties suggest that the constituent ECM molecules released from the gel remain activity. These findings suggested that the SIS gel was a promising candidate for tissue engineering, especially in promoting vessel formation.

Generating Elastin-Rich Small Intestinal Submucosa–Based Smooth Muscle Constructs Utilizing Exogenous Growth Factors and Cyclic Mechanical Stimulation

Successful approaches to tissue engineering smooth muscle tissues utilize biodegradable scaffolds seeded with autologous cells. One common problem in using biological scaffolds specifically is the difficulty of inducing cellular penetration and controlling de novo extracellular matrix deposition/remodeling in vitro. Our hypothesis was that small intestinal submucosa (SIS) exposed to specific mechanical stimulation regimes would modulate the synthesis of de novo collagen and elastin by bladder smooth muscle cells (BSMC) within the SIS matrix. We further hypothesized that the cytokines vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF-2), two key growth factors involved in epithelial mesenchymal signaling, will promote the cellular penetration into SIS necessary for mechanical stimulation. BSMC were seeded at 0.5 x 10(6) cells/cm(2) onto the luminal side of SIS specimens. VEGF (10 ng/mL) and FGF-2 (5 ng/mL) were added to each insert in the media every other day for up to 7 days in static culture. Following static culture, specimens were stretched strip-biaxially under 15% peak strain at either 0.5 or 0.1 Hz for an additional 7 days. Following the culture period, specimens were assayed histologically and biochemically for cellular penetration, proliferation, elastin, collagen, and protease activity. Histological analyses demonstrated that in standard culture media, BSMC remained on the surface of the SIS while both FGF-2 and VEGF profoundly promoted ingrowth of the BSMC into the SIS. The penetration of the cells in response to these cytokines was confirmed using a Transwell assay. Following cellular penetration, BSMC produced significant amounts of elastic fibers under cyclic mechanical stretching at 0.1 Hz under 15% stretch, as evidenced by colorimetric assay and histology using a Verhoeff-Van Gieson stain. Protease activity was assessed in the media and found to be statistically increased in static culture following FGF-2 treatment. These findings demonstrate, for the first time, the capability of BSMC to produce histologically apparent elastin fibers in vitro. Moreover, our results suggest that a strategy involving growth factors and controlled mechanical stimulation may be used to engineer functional, elastin-rich tissue replacements using decellularized biologically derived scaffolds.

Effects of sterilization on an extracellular matrix scaffold: Part II. Bioactivity and matrix interaction

Small intestinal submucosa (SIS) has been successfully used to treat a variety of damaged or diseased tissues in human patients. As a biologic scaffold, SIS stimulates repair of damaged or diseased tissues and organs with tissue that is similar in structure and function to the material it was meant to replace. To meet clinical safety requirements, biologic materials from animal tissues must undergo processing treatments to minimize host immune response and to eliminate the possibility of disease transmission. The effect of peracetic acid disinfection, lyophilization, and ethylene oxide sterilization on the in vitro bioactivity of the processed SIS was therefore examined in murine fibroblasts and pheochromocytoma (PC12) cells. Specifically, the ability of processed SIS to support fibroblast attachment, to stimulate PC12 cell differentiation, and to upregulate fibroblast VEGF secretion was examined. Fibroblasts attach to the sterilized SIS, remain viable, and more than double their secretion of VEGF as a result of interacting with the SIS matrix components. Additionally, PC12 cells exhibit increased neurite outgrowth following stimulation by SIS matrix proteins versus controls. We conclude that a biologic scaffold can be prepared for human use and still retain significant bioactivity.

Porcine Small Intestinal Submucosa Graft for Repair of Anterior Urethral Strictures

ObjectivesWe evaluated porcine small intestinal submucosa (SIS) used in the treatment of inflammatory, iatrogenic, posttraumatic, and idiopathic strictures of bulbar and penile urethra. Midterm maintenance of urethral patency was assessed. MethodsFifty patients aged 45–73 yr with anterior urethral stricture underwent urethroplasty using a porcine SIS collagen-based matrix for urethral reconstruction. Stricture was localized in the bulbar urethra in 10 patients, the bulbopenile area in 31 cases, and in the distal penile urethra in nine patients. All patients received a four-layered SIS patch graft in an onlay fashion. A voiding history, retrograde and antegrade urethrography, and cystoscopy were performed preoperatively and postoperatively. Failure was defined as stricture confirmed on urethrogram. ResultsAfter a mean follow-up of 31.2 mo (range: 24–36 mo), the clinical, radiological, and cosmetic findings were excellent in 40 (80%) patients. Restricture developed in one of 10 bulbar, five of 31 bulbopenile, and four of nine penile strictures. These all occurred in the first 6 mo postoperatively. All patients with recurrences needed further therapy, but there has been no additional recurrence observed to date. No complications such as fistula, wound infection, UTI, or rejection were observed. ConclusionsUse of inert porcine SIS matrix appears to be beneficial for patients with bulbar and bulbopenile strictures. Midterm results are comparable to skin flaps and mucosal grafts.

USE OF SMALL INTESTINAL SUBMUCOSE FOR TISSUE REGENERATION IN MAXILLOFACIAL SURGERY A SAFETY STUDY

Small intestinal submucosa (SIS) is a promising material for guided tissue regeneration (CTR). Clinical and in vitro experiments within other fields of regenerative surgery had shown high efficiency and biocompatibility. No clinical trials had been developed for oral reconstructive surgery. The objective of this study was to evaluate the safety of new standardized SIS production method as a biomaterial for GTR in maxillofacial surgery. Methods: SIS membranes were produced by a standardized method developed by our research group. Patients with oral defects requiring bone grafting were recruited and treated using autologous bone grafts and SIS membranes. Informed consent was obtained from all patients. Clinic and radiographic data were recorded for each patient pre and post operatively. In vitro cells culture was performed using SIS matrix and fibroblasts prior clinical use to measure biocompatibility. Immune systemic response was evaluated through acute systemic toxicity, pyrogenicity and allergic tests. Results: 49 patients were treated. Age ranged between 21 and 74 years. 37 patients had atrophic bone defects, 7 had infectious bone defects, 2 had traumatic bone defects and 1 had bone defect after mechanical distractor device placement. Optimal clinical and radiographic results were found in response to the surgical treatment. Only two patients showed complications related with dental implants placement after CTR but these complications could not be associated with the use of SIS. As a conclusion SIS showed to be a highly biocornpatible material, adverse reactions to SIS were not found in any case.

Small intestinal submucosa: a substrate for in vitro cell growth

Abstraet-The extracellular matrix (ECM) of the small intestinal submucosa (SIS) was harvested by removing the superficial layers of the mucosa and the external muscular layers. The remaining 80 μm thick sheet was disinfected and sterilized by methods which removed all cellular components. The SIS-ECM, retaining its native 3-dimensional microarchitecture and composition, was evaluated for its ability to support in vitro cell growth. Six separate cell types were seeded either alone or in coculture with other cells upon this matrix, grown in selected media, and examined daily for time periods ranging from 48 h to 2 weeks. The six cell types tested were NIH Swiss mouse 3T3 fibroblasts, NIH 3T3/J2 fibroblasts, primary human fibroblasts, primary human keratinocytes, human microvascular endothelial cells (HMECs), and an established rat osteosarcoma (ROS) cell line. All cell types showed the ability to attach and proliferate. All fibroblast cell lines and the keratinocytes proliferated and/or migrated into the 3-dimensional scaffold of the SIS matrix. The ROS cells and the HMECs were confined in their growth pattern to the surface of the matrix. Coculturing of NIH 3T3/J2 fibroblasts and primary human keratinocytes resulted in a distinctive spatial orientation of the two cell types. The fibroblasts populated the mid-substance of the 3-dimensional matrix and the keratinocytes formed an epidermal structure with rete ridge-like formation and stratification when the composite was lifted to an air liquid interface in culture. In summary, SIS provides a substratum with a 3-dimensional scaffold that allows for cell migration and spatial organization. This substratum is suitable for in vitro studies of the interaction between epithelial or mesenchymal cells and a naturally occurring extracellular matrix.