In situ forming gelatin/hyaluronic acid hydrogel for tissue sealing and hemostasis.

Fibrin glue has been widely used as a surgical sealing and hemostatic agent. Its application is restricted due to poor tissue adhesion and low mechanical strength. To develop better tissue sealant and hemostatic agent, this study prepared the injectable hydrogels by chemically cross-linking gelatin (G) with or without hyaluronic acid (HA) in situ at a mild condition. The rheological analysis, Fourier transform infrared spectroscopy, swelling, proteolytic degradation, biocompatibility, tissue sealing, and hemostatic ability of the hydrogels were investigated. It was found that the chemical cross-linking rapidly formed in both self-crosslinking gelatin (sc-G) and gelatin/hyaluronate acid (G/HA) hydrogels. The hydrogels could be degraded by trypsin and had a desirable biocompatibility. The tissue sealing ability of the hydrogels was superior to fibrin glue. Furthermore, the G/HA hydrogel had similar hemostatic performance as fibrin glue, and was better than that of gelatin hydrogel. The results in the study indicated that the G/HA hydrogel could be used in clinic as a tissue sealant or surgical hemostat.

In situ injectable hyaluronic acid/gelatin hydrogel for hemorrhage control.

Tissue sealants are used for hemorrhage control which is imperative in many surgical procedures. It is a highly challenging task to obtain the ideal tissue sealant. Only a few commercially tissue sealants are available to be used for internal tissue or organ hemorrhage control. This study introduced two in situ injectable hydrogels for hemorrhage control: self-crosslinking gelatin (sc-G) hydrogel and hyaluronic acid/gelatin (HA/G) hydrogel. They were prepared on the tissue surface in situ and characterized by rheological analysis, stability, cytotoxicity, and bursting strength test. The hemostatic ability of the hydrogels was evaluated in a liver-bleeding rat model. The sc-G and HA/G hydrogels gelled around 90 s and 50 s, respectively. They were preferable for cell attachment and proliferation. The bursting strengths of both hydrogels exceeded that of fibrin glue. The hemostatic ability of HA/G hydrogel was better than that of sc-G hydrogel, and was same as that of fibrin glue. The HA/G hydrogel could be used as a tissue sealant for hemorrhage control in clinic.

Tendon-Derived Stem Cell Differentiation in the Degenerative Tendon Microenvironment.

Tendinopathy is prevalent in athletic and many occupational populations; nevertheless, the pathogenesis of tendinopathy remains unclear. Tendon-derived stem cells (TDSCs) were regarded as the key culprit for the development of tendinopathy. However, it is uncertain how TDSCs differentiate into adipocytes, chondrocytes, or osteocytes in the degenerative microenvironment of tendinopathy. So in this study, the regulating effects of the degenerative tendon microenvironment on differentiation of TDSCs were investigated. TDSCs were isolated from rat Achilles tendons and were grown on normal and degenerative (prepared by stress-deprived culture) decellularized tendon slices (DTSs). Immunofluorescence staining, H&E staining, real-time PCR, and Western blot were used to delineate the morphology, proliferation, and differentiation of TDSCs in the degenerative microenvironment. It was found that TDSCs were much more spread on the degenerative DTSs than those on normal DTSs. The tenocyte-related markers, COL1 and TNMD, were highly expressed on normal DTSs than the degenerative DTSs. The expression of chondrogenic and osteogenic markers, COL2, SOX9, Runx2, and ALP, was higher on the degenerative DTSs compared with TDSCs on normal DTSs. Furthermore, phosphorylated FAK and ERK1/2 were reduced on degenerative DTSs. In conclusion, this study found that the degenerative tendon microenvironment induced TDSCs to differentiate into chondrogenic and osteogenic lineages. It could be attributed to the cell morphology changes and reduced FAK and ERK1/2 activation in the degenerative microenvironment of tendinopathy.

Matrix stiffness regulates the differentiation of tendon-derived stem cells through FAK-ERK1/2 activation.

Tendon derived stem cells (TDSCs) were vital in tendon homeostasis. Nevertheless, the regulation of TDSCs differentiation in tendinopathy is unclear. Matrix stiffness modulated stem cells differentiation, and matrix stiffness of tendinopathic tissues decreased significantly. In order to clarify the role of matrix stiffness in TDSCs differentiation, they were cultured on the gelatin hydrogels with the stiffness from 2.34 ±â€¯1.48 kPa to 24.09 ±â€¯14.03 kPa. The effect of matrix stiffness on TDSCs proliferation and differentiation were investigated with CCK8 assay, immunofluorescences, real time PCR and western blot. It was found the proliferation of TDSCs increased and more stress fibers formed with increasing matrix stiffness. The differentiation of TDSCs into tenogenic, chondrogenic, and osteogenic lineages were inhibited on stiff hydrogel evidenced by reduced expression of tenocyte markers THBS4, TNMD, SCX, chondrocyte marker COL2, and osteocyte markers Runx2, Osterix, and ALP. Furthermore, the phosphorylation of FAK and ERK1/2 were enhanced when TDSCs grew on stiff hydrogel. After FAK or ERK1/2 was inhibited, the effect of matrix stiffness on differentiation of TDSCs was inhibited as well. The above results indicated matrix stiffness modulated the proliferation and differentiation of TDSCs, and the regulation effect could correlate to the activation of FAK or ERK1/2.

In situ cross-linking carbodiimide-modified chitosan hydrogel for postoperative adhesion prevention in a rat model.

BACKGROUND: Postoperative intra-abdominal adhesion often causes many complications. Chitosan fluid has been used in clinic to prevent intra-abdominal adhesion. However, fluid can be easily diluted and cannot stay on the wound site. As hydrogel is able to form stable physical barrier to separate injured tissues, we developed a chitosan hydrogel for better prevention of intra-abdominal adhesion in this study. METHODS: We synthesized a carbodiimide-derivatized chitosan gelatin (cd-CS-gelatin) hydrogel and investigated its rheological properties. A rat model was used to compare the anti-adhesion effect of chitosan hydrogel and fluid. The wounds were created with damage of the underlying muscle of the abdominal wall and the serosal layer of the cecum. They were coated with chitosan fluid or cd-CS-gelatin hydrogel. At day 14 after surgery, the animals were euthanized and intra-abdominal adhesion was assessed. RESULTS: The cd-CS-gelatin hydrogel solidified within 3min after the mixing of the reagents. The cecum-abdomen adhesion occurred in all rats without anti-adhesion treatment. The application of cd-CS-gelatin significantly reduced the adhesion rate from 100% to 50%, compared the chitosan fluid only to 88%. The decrease of adhesion breaking strength also manifested that cd-CS-gelatin was more effective than chitosan fluid to reduce postsurgical intra-abdominal adhesion formation. CONCLUSIONS: Chitosan hydrogel is more effective than chitosan fluid to prevent postoperative cecum-abdomen adhesion. It indicates that hydrogel could be a more promising state than liquid to prevent postoperative intra-abdominal adhesion.

Evaluation of surgical anti-adhesion products to reduce postsurgical intra-abdominal adhesion formation in a rat model.

BACKGROUND: Adhesions frequently occur after abdominal surgery. Many anti-adhesion products have been used in clinic. However, the evidences are short for surgeons to reasonably choose the suitable anti-adhesion produces in clinical practice. This study provided such evidence by comparing the efficiency of five products to prevent abdominal adhesion formation in a rat model. METHODS: Fifty-six Sprague-Dawley rats were randomly divided into seven groups: sham-operation group, adhesion group, and five product groups (n = 8). The abdomens of rats were opened. The injuries were created on abdominal wall and cecum in the adhesion and product groups. The wounds on abdominal wall and cecum of rats in the adhesion group were not treated before the abdomens were closed. The wounds on abdominal wall and cecum of rats in the product groups were covered with anti-adhesion product: polylactic acid (PLA) film, Seprafilm®, medical polyethylene glycol berberine liquid (PEG), medical sodium hyaluronate gel (HA), or medical chitosan (Chitosan). Fourteen days after surgery, the adhesions were evaluated by incidence, severity, adhesion area on abdominal wall and adhesion breaking strength. RESULTS: The application of PLA film and Seprafilm® significantly reduced the incidence, severity, adhesion area and breaking strength of cecum-abdomen adhesion (P<0.05). HA, PEG and Chitosan failed to significantly reduce the cecum-abdomen adhesion (P>0.05). The statistical significances in the incidence and severity of abdomen-adipose adhesion between adhesion group and the product groups were not achieved. However, Seprafilm® was more effective to reduce abdomen-adipose adhesion than PLA film. Furthermore, it was found that the products tested in this study did not effectively reduce cecum-adipose adhesion. The application of PEG could result in abdomen-small intestine adhesion. CONCLUSION: Based on the results of this study, the preference order of anti-adhesion products used to reduce postsurgical intra-abdominal adhesion formation is Seprafilm > PLA >> HA > Chitosan > PEG.

Work of separation - A method to assess intraperitoneal adhesion and healing of parietal peritoneum in an animal model.

BACKGROUND: Adhesion grades and adhesion breaking strength are widely used to assess severity of intraperitoneal adhesion in animal models. However, the results of adhesion grades have the large deviations due to vary personal experience. Adhesion breaking strength ignores the details of adhesion. This study introduced work of separation, the energy consumption during breakage of adhesion, to better evaluate intraperitoneal adhesion. METHODS: The intraperitoneal adhesion was induced by traumas created at rat cecum and adjacent abdominal wall. The wounds were coated with or without sodium hyaluronate. On day 14 after surgery, the intraperitoneal adhesion was assessed by adhesion density grade, adhesion area grade, adhesion breaking strength and work of separation. The healing of parietal peritoneum was evaluated with histology, adhesion breaking strength and work of separation. FINDINGS: The severity of adhesion evaluated with work of separation was consistent with those obtained from the grades of adhesion density, adhesion area and adhesion breaking strength. Work of separation had a linear correlation with adhesion breaking strength. Furthermore, the results of histological examination and work of separation demonstrated that adhesion significantly delayed healing process of abdominal wall muscles. INTERPRETATION: Work of separation can quantify all intraperitoneal adhesions rather than the major one by other methods. It is a more precise method to evaluate postoperative adhesions, especially those including adipose tissue. This study proved that work of separation could be a reliable method to assess intraperitoneal adhesion and tissue healing.

Effect of carbodiimide-derivatized hyaluronic acid gelatin on preventing postsurgical intra-abdominal adhesion formation and promoting healing in a rat model.

Adhesions often occur after abdominal surgery. It could cause chronic pelvic pain, intestinal obstruction, and infertility. A hydrogel biomaterial, carbodiimide-derivatized hyaluronic acid gelatin (cd-HA gelatin), has been successfully used to reduce adhesion formation after flexor tendon grafting. This study investigated the efficacy of cd-HA gelatin in preventing postsurgical peritoneal adhesions in a rat model. The surgical traumas were created on the underlying muscle of the abdominal wall and the serosal layer of the cecum. The wounds were covered with or without cd-HA gelatin. Animals were euthanized at day 14 after surgery. Adhesion formation was assessed with adhesion degree and adhesion breaking strength. The healing of abdominal wall was evaluated with biomechanical testing and histological analysis. The adhesions occurred in all rats (n = 12) without cd-HA gelatin treatment. The application of cd-HA gelatin significantly reduced the adhesion rate from 100% to 58%. The decrease of adhesion breaking strength also manifested that cd-HA gelatin could reduce postsurgical intra-abdominal adhesion formation. Moreover, it was found that cd-HA gelatin was a safe material and could promote tissue healing. The cd-HA gelatin hydrogel could reduce the formation of intra-abdominal adhesions without adversely effects on wound healing.

Anodic TiO2 nanotubular arrays with pre-synthesized hydroxyapatite--an effective approach to enhance the biocompatibility of titanium.

Electrochemically anodized TiO2 nanotubular arrays can provide large surface areas for biological species attachment. In order to further enhance the biocompatibility of Ti medical implants, we deposited a pre-synthesized hydroxyapatite inside and on the nanotubular arrays, and examined the biocompatibility of the anodized TiO2 nanotubular arrays with pre-synthesized hydroxyapatite by in vitro assessment in simulated body fluid, and in vitro cell culture. The results showed that the hydroxyapatite coating was able to be induced on TiO2 nanotubular arrays with pre-synthesized hydroxyapatite within 5 days while only a thin film composed of calcium phosphorous chemicals formed on as-formed TiO2 nanotubular arrays. The cell culture evaluation further proved the enhancement of cell attachment and proliferation on TiO2 nanotubular arrays with pre-synthesized hydroxyapatite as opposed to those without pre-synthesized hydroxyapatite. The present study proves that formation of TiO2 nanotubular arrays with pre-synthesized hydroxyapatite a promising method to enhance the biocompatibility of Ti implants.