Experimental Correction and Treatment of Chronic Liver Failure Using Implantable Cell-Engineering Constructs of the Auxiliary Liver Based on a Bioactive Heterogeneous Biopolymer Hydrogel.

Our study sought approaches for chronic liver failure (CLF) treatment and correction via cell-engineered constructs (CECs). They are built from biopolymer-based, microstructured, and collagen-containing hydrogel (BMCG). We also strove to evaluate the functional activity of BMCG in liver regeneration. MATERIALS AND METHODS: Allogeneic liver cells (namely, hepatocytes; LC) together with mesenchymal multipotent stem cells of bone marrow origin (MMSC BM; BMSCs) were adhered to our BMCG to compose implanted liver CECs. Thereafter, we investigated a model of CLF in rats receiving the implanted CECs. The CLF had been provoked by long-term exposure to carbon tetrachloride. The study comprised male Wistar rats (n = 120) randomized into 3 groups: Group 1 was a control group with the saline treatment of the hepatic parenchyma (n = 40); Group 2 received BMCG only (n = 40); and Group 3 was loaded with CECs implanted into the parenchyma of their livers (n = 40). August rats (n = 30) made up a donor population for LCs and MMSC BM to develop grafts for animals from Group 3. The study length was 90 days. RESULTS: CECs were shown to affect both biochemical test values and morphological parameters in rats with CLF. CONCLUSION: We found BMCG-derived CECs to be operational and active, with regenerative potential. Group 3 showed significant evidence of forced liver regeneration that tended to persist until the end of the study (day 90). The phenomenon is reflected by biochemical signs of hepatic functional recovery by day 30 after grafting (compared to Groups 1 and 2), whereas structural features of liver repair (necrosis prevention, missing formation of vacuoles, degenerating LC number decrease, and delay of hepatic fibrotic transformation). Such implantation of BMCG-derived CECs with allogeneic LCs and MMSC BM might represent a proper option to correct and treat CLF, as well as to maintain affected liver function in patients with liver grafting needed.

Decellularization of cartilage microparticles: Effects of temperature, supercritical carbon dioxide and ultrasound on biochemical, mechanical, and biological properties.

One of the approaches to restoring the structure of damaged cartilage tissue is an intra-articular injection of tissue-engineered medical products (TEMPs) consisting of biocompatible matrices loaded with cells. The most interesting are the absorbable matrices from decellularized tissues, provided that the cellular material is completely removed from them with the maximum possible preservation of the structure and composition of the natural extracellular matrix. The present study investigated the mechanical, biochemical, and biological properties of decellularized porcine cartilage microparticles (DCMps) obtained by techniques, differing only in physical treatments, such as freeze-thaw cycling (Protocol 1), supercritical carbon dioxide fluid (Protocol 2) and ultrasound (Protocol 3). Full tissue decellularization was achieved, as confirmed by the histological analysis and DNA quantification, though all the resultant DCMps had reduced glycosaminoglycans (GAGs) and collagen. The elastic modulus of all DCMp samples was also significantly reduced. Most notably, DCMps prepared with Protocol 3 significantly outperformed other samples in viability and the chondroinduction of the human adipose-derived stem cells (hADSCs), with a higher GAG production per DNA content. A positive ECM staining for type II collagen was also detected only in cartilage-like structures based on ultrasound-treated DCMps. The biocompatibility of a xenogenic DCMps obtained with Protocol 3 has been confirmed for a 6-month implantation in the thigh muscle tissue of mature rats (n = 18). Overall, the results showed that the porcine cartilage microparticles decellularized by a combination of detergents, ultrasound and DNase could be a promising source of scaffolds for TEMPs for cartilage reconstruction.

Upregulated circulating mir-424 and its' diagnostic value for gram-negative bacteremia after thoracic transplantation.

Aims: Early post-transplant complications such as acute graft rejection and infections are associated with high morbidity and mortality of heart and lung transplant recipients who are in vital need of immunosuppressive therapy. MiR-424 is a member of the miR-16 family, which plays an important physiological role in the development of cardiovascular and respiratory pathology, is involved in the regulation of monocyte and macrophage differentiation, and has an immunosuppressive potential. The aim of the study was to determine the diagnostic value of circulating miR-424 as a potential biomarker of post-transplant complications in heart and lung transplant recipients. Methods: The study enrolled 83 heart transplant recipients, aged 18 to 70 (48 ± 13) years; 26 lung transplant recipients, aged 10 to 74 (36 ± 16) years. The miR-424 plasma expression was detected by real-time PCR (Qiagen, USA). Significance of miR-424 level was assessed through the ΔCt method. Acute graft rejection was verified by the results of endomyocardial or transbronchial biopsy. Post-transplant infectious complications were verified through microbiological identification of bacteremia from blood cultures. Results: Our study shows miR-424 upregulation in plasma of patients with chronic heart or respiratory failure in comparison with healthy individuals (p = 0.003 and p = 0.04 resp.). There was a direct correlation of miR-424 expression with red blood cells and hemoglobin levels in patients before heart transplantation (p = 0.01 and p = 0.03 resp.). After transplantation the expression of plasma miR-424 correlated with the level of C-reactive protein (CRP) both in heart (r = 0.75; p = 0.02) and lung (r = 0.50; p = 0.04) transplant recipients. The expression of plasma miR-424 correlated with tacrolimus blood concentration after heart transplantation (r = 0.38; p = 0.04). The miR-424 level didn't differ in heart or lung transplant recipients with and without acute graft rejection (p = 0.47 and p = 0.78 resp.), but was significantly higher in heart and lung transplant recipients with gram-negative bacteremia (p = 0.002). When the miR-424 level is above a threshold value (-5.72 fold change), the relative risk of bacteremia is RR = 3.84 [95% CI 1.94-7.61]; Se = 60.0%; Sp = 89.2%. CRP concentration above 7 mg/L in duplex test with miR-424 improves the diagnostic characteristics of miR-424 for post-transplant gram-negative bacteremia in heart and lung transplant recipients up to RR = 9.17 [95% CI 1.37-61.46]; Se = 83.3% and Sp = 90.1%. Conclusion: MiR-424 plasma expression was upregulated in patients with chronic heart and respiratory failure and in heart and lung transplant recipients in the early post-transplant period. The duplex test, including miR-424 and CRP, has a diagnostic value for detecting the high risk of post-transplant gram-negative bacteremia in heart and lung transplant recipients.

Recombinant Spidroin Microgel as the Base of Cell-Engineered Constructs Mediates Liver Regeneration in Rats.

Aim: In this study, we seek to check if recombinant spidroin rS1/9 is applicable for cell-engineering construct development. Novel technologies of cell and tissue engineering are relevant for chronic liver failure management. Liver regeneration may represent one of the possible treatment options if a cell-engineered construct (CEC) is used. Nowadays, one can see the continuous study of various matrices to create an appropriate CEC. Materials and Methods: We have adhered allogenic liver cells and multipotent mesenchymal bone marrow stem cells (MMSC BM) to a microgel with recombinant spidroin rS1/9. Then we have studied the developed implantable CEC in a rat model (n = 80) of chronic liver failure achieved by prolonged poisoning with carbon tetrachloride. Results: Our results demonstrate that the CECs change the values of biochemical tests and morphological parameters in chronic liver failure in rats. Conclusion: We consider there to be a positive effect from the microgel-based CECs with recombinant spidroin rS1/9 in the treatment of chronic liver failure.

Decellularization of Human Pancreatic Fragments with Pronounced Signs of Structural Changes.

A significant lack of donor organs restricts the opportunity to obtain tissue-specific scaffolds for tissue-engineering technologies. One of the acceptable solutions is the development of decellularization protocols for a human donor pancreas unsuitable for transplantation. A protocol of obtaining a biocompatible tissue-specific scaffold from decellularized fragments with pronounced human pancreas lipomatosis signs with preserved basic fibrillary proteins of a pancreatic tissue extracellular matrix was developed. The scaffold supports the adhesion and proliferation of human adipose derived stem cell (hADSCs) and prolongs the viability and insulin-producing function of pancreatic islets. Experiments conducted allow for the reliance on the prospects of using the donor pancreas unsuitable for transplantation in the technologies of tissue engineering and regenerative medicine, including the development of a tissue equivalent of a pancreas.

MiR-339 and galectin-3: diagnostic value in patients with airway obstruction after lung transplantation.

Respiratory complications can be the cause of graft dysfunction after lung transplantation (LTx). MicroRNAs are small regulatory molecules-potential biomarkers of respiratory diseases and post-transplant complications. Galectin-3 is highly expressed in fibrosis of transplanted solid organs. The aim was to evaluate the expression of plasma miR-339 and galectin-3 concentrations in lung recipients including with airway obstruction after LTx. The study included 57 lung recipients (34 men and 23 women aged 10 to 74 years) were followed up to 5 years after LTx. The plasma microRNAs were detected by real-time PCR; galectin-3 levels were measured by ELISA. During follow-up in 30 recipients, post-transplant complications were detected: 12 (40.0%) cases of airway obstruction. The levels of miR-339 and galectin-3 were significantly higher in recipients with airway obstruction compare with 27 (47.3%) recipients without any complications (P = 0.036 and P = 0.014, resp.). Increasing miR-339 (above the 0.02 fold change) and galectin-3 (above the 11.7 ng/ml) threshold plasma levels in lung recipients is associated with high risk (RR = 7.14 ± 0.97 [95% CI 1.05-48.60], P = 0.045) of airway obstruction after LTx. A measurement of miR-339 expression in combination with galectin-3 level might be perspective to identify recipients at risk of airway obstruction after LTx.

A Comparison of the Capacity of Mesenchymal Stromal Cells for Cartilage Regeneration Depending on Collagen-Based Injectable Biomimetic Scaffold Type.

Mesenchymal stromal cells (MSCs) have shown a high potential for cartilage repair. Collagen-based scaffolds are used to deliver and retain cells at the site of cartilage damage. The aim of the work was a comparative analysis of the capacity of the MSCs from human adipose tissue to differentiate into chondrocytes in vitro and to stimulate the regeneration of articular cartilage in an experimental model of rabbit knee osteoarthrosis when cultured on microheterogenic collagen-based hydrogel (MCH) and the microparticles of decellularized porcine articular cartilage (DPC). The morphology of samples was evaluated using scanning electron microscopy and histological staining methods. On the surface of the DPC, the cells were distributed more uniformly than on the MCH surface. On day 28, the cells cultured on the DPC produced glycosaminoglycans more intensely compared to the MCH with the synthesis of collagen type II. However, in the experimental model of osteoarthrosis, the stimulation of the cartilage regeneration was more effective when the MSCs were administered to the MCH carrier. The present study demonstrates the way to regulate the action of the MSCs in the area of cartilage regeneration: the MCH is more conducive to stimulating cartilage repair by the MSCs, while the DPC is an inducer for a formation of a cartilage-like tissue by the MSCs in vitro.

Time is of the essence: A single-center experience of hepatic arterial supply impairment management in pediatric liver transplant recipients.

BACKGROUND: Impairment of hepatic arterial flow including hepatic arterial thrombosis (HAT), hepatic arterial stenosis (HAS), and splenic artery steal syndrome (SASS) is potentially life-threatening complications. The proposed early diagnosis and urgent treatment strategy of graft arterial flow reduction aim to decrease morbidity and mortality. METHODS: Pediatric patients with known hepatic arterial flow impairment were retrospectively reviewed. Patients were grouped by occlusive (HAT) and non-occlusive (HAS/SASS) arterial flow reduction. Patients with HAT were further divided in two groups based on the estimated maximal hepatic artery occlusion time ≤8 and >8 hours. RESULTS: Impairment of hepatic arterial flow developed in 32 of 416 pediatric liver transplant recipients. HAT, HAS, and SASS incidences were 4.1% (n = 17), 2.2% (n = 9), and 1.4% (n = 6), respectively. Neither graft loss nor death occurred in the non-occlusive group. The probabilities of sepsis (OR, 1.7; 95% CI, 1.14-2.53; P=.008) and graft loss or death (OR, 1.42; 95% CI, 1.04-1.92; P=.046) were higher in the occlusive group. Patients with estimated maximal duration of hepatic artery occlusion ≤ 8 hours (n = 7; 41.2%) did not have ischemic-type biliary lesions and sepsis (P=.044 and 0.010, respectively) but had excellent 3-year graft survival compared with > 8 hours group (100% vs 40%; P=.037). Multivariate analysis revealed HAT manifestation by fever was associated with increased chances of graft loss or death. CONCLUSION: Occlusive arterial complications impose higher risks of graft loss and death. Thorough arterial supply monitoring by Doppler ultrasonography and urgent endovascular arterial flow restoration may salvage both graft and the recipient.

Transforming growth factor beta 1 levels in the blood of pediatric liver recipients: Clinical and biochemical correlations.

TGF-ß1 is a cytokine with profibrogenic and immunosuppressive activities, which suggest the clinical significance of TGF-ß1 for the assessment of graft function after LT. We analyzed the dynamics of TGF-ß1 levels in the blood after LDLT in 135 pediatric liver recipients and examined the relationship between the cytokine levels and the laboratory and clinical variables. We found that TGF-ß1 levels in the blood of patients with ESLD were lower than that in healthy children of the same age, P = .001. Moreover, blood levels of TGF-ß1 were associated with liver disease etiology (r = .23) and hepatic fibrosis severity (r = .33). Before LDLT, TGF-ß1 levels were significantly higher in children with good outcomes than in recipients who developed graft dysfunction early in the post-transplant period, P = .047. One month after LDLT, TGF-ß1 levels in blood plasma increased in pediatric recipients, P = .002. Cytokine levels were significantly correlated with gender (r = .21) and HLA (r = -.24) mismatches, as well as with TAC dosage (r = -.32) later in the post-transplant period. One year after LDLT, TGF-ß1 plasma levels were higher (P = .01) than those before LDLT and did not correlate with most of the investigated biochemical and clinical variables. Conclusion: Blood levels of TGF-ß1 are associated with hepatic fibrosis severity, graft dysfunction development, and TAC dosage and can be regarded as a potential prognostic biomarker for the assessment of graft function and the optimization of immunosuppressant dosage in pediatric recipients after LDLT.

Deceased vs living donor grafts for pediatric simultaneous liver-kidney transplantation: A single-center experience.

INTRODUCTION: In conditions of limited experience of pediatric simultaneous liver-kidney transplantation (SLKT) using grafts from living and deceased donors, there is a certain need to validate the approach. PATIENTS: The retrospective study of 18 pediatric patients who received SLKT between 2008 and 2019. RESULTS: Grafts were obtained from both living and deceased donors. The patients' age ranged from 2 to 16 years (9 years ±4). The body weight of the children varied from 9.5 to 39 kg (22 kg ±9). The follow-up period lasted from 1 to 109 months (median 38 months ±35). The various graft combinations were used in both groups. There was no mortality during the follow-up. There was no significant difference in baseline parameters in recipients who received grafts from living and deceased donors except age (7.5 years ±2.2 vs 11.8 years ±4.1; P = .038). Rate of complications > grade II was higher among recipients of deceased donor SLKT (7.7% vs 60%; OR, 7.8; 95% CI, 1.04-58.48; P = .044). All the patients are alive with both grafts functioning. All the living donors returned to the normal life. CONCLUSION: SLKT is a safe and effective procedure for children with both simultaneous end-stage liver disease and end-stage renal disease. Both living donor partial liver and kidney transplantation and deceased donor liver-kidney transplantation can be considered as safe and feasible options.

Simultaneous laparoscopic left lateral sectionectomy and nephrectomy in the same living donor: The first case report.

With the presence of organ shortage, living donors remain important sources of grafts, especially for pediatric recipients. Laparoscopic nephrectomy has become the gold standard for living donors. Additionally, laparoscopic partial liver procurement in living donors has proven its safety and feasibility in the latest studies. We have combined both approaches to perform a simultaneous liver-kidney transplantation in a pediatric patient from the same living donor. Our experience of laparoscopic left lateral sectionectomy and laparoscopic nephrectomy in living donors was the basis for adapting to this procedure. A 29-year-old mother was an ABO-incompatible (ABOi) donor for the left lateral section (LLS) of the liver and left kidney for her 2-year-old son. The postoperative period was uneventful. Two sessions of plasmapheresis and rituximab induction were necessary to prepare for ABOi transplantation. The donor and recipient were discharged on postoperative days 5 and 28, respectively. Simultaneous laparoscopic left lateral sectionectomy and nephrectomy in the same living donor is feasible for transplantation from the parent to the child with advanced laparoscopic expertise.

Laparoscopic left lateral section procurement in living liver donors: A single center propensity score-matched study.

BACKGROUND: Laparoscopic living donor liver procurement for transplantation has increased in popularity over the past decade. The purpose of this study was to compare the laparoscopic and open approaches in living donor left lateral sectionectomy (LLS) and to assess the safety and feasibility of this laparoscopic approach. METHODS: A total of 103 living donor LLSs were performed at our center from May 2016 to December 2017. Of these, 35 were completely laparoscopic procedures, which represented the subject of this study. An additional 68 open living donor LLSs performed during the same period were studied as a comparison group. To overcome selection bias, LLS donors were balanced on a 1:1 ratio (laparoscopic [n = 35]: open [n = 35]) according to covariates with similar values. The PSM was based on the operation date, recipient age, diagnosis, recipient weight, and donor age. RESULTS: There were significant differences between the laparoscopic and open LLS groups (P < 0.001) in terms of blood loss (96.8 ± 16.5 vs 155.8 ± 17.8 mL) as well as the duration of hospital stay (4 ± 0.4 vs 6.9 ± 0.5 days). CONCLUSION: Laparoscopic LLS is a feasible and efficacious in the setting of a developed program with advanced laparoscopic expertise.

The hematopoietic stem cell number in the peripheral blood of pediatric recipients correlates with the outcome after living donor liver transplantation.

It has been proposed that circulating HSCs play a role in graft survival after liver transplantation. The aim was to analyze the relationship between the number of HSCs before and after LDLT and liver function, immune biomarkers, and clinical outcomes in pediatric patients. We studied 15 pairs of adult healthy liver donors and pediatric recipients with ESLD. The CD34/CD45+ cell number was measured in the blood via flow cytometry, and plasma levels of immune biomarkers - via ELISA. CD34/CD45+ cell number in the recipients decreased within the first week after LDLT. The cell number before LDLT was negatively correlated with the plasma levels of CRP and the development of graft dysfunction in the early post-transplant period. After LDLT, the CD34/CD45+ cell number was positively correlated with the pretransplant plasma level of sCD40L, a T-cell activation marker. In adult liver donors, the cell number did not change within the first week after liver resection and was lower than in pediatric recipients. The results suggest that in pediatric recipients, the HSC number may be associated with graft function and could be regarded as a potential predictor of the clinical outcome after LDLT.