Flavia Neri

Mesenchymal Stem Cells and Islet Cotransplantation in Diabetic Rats: Improved Islet Graft Revascularization and Function by Human Adipose Tissue-Derived Stem Cells Preconditioned With Natural Molecules

Hypoxia plays an important role in limiting the engraftment, survival, and function of intrahepatically transplanted islets. Mesenchymal stem cells (MSCs) were recently used in animal models of islet transplantation not only to reduce allograft rejection but also to promote revascularization. Among different possible origins, adipose tissue represents a novel and good source of MSCs. Moreover, the capability of adipose tissue-derived stem cells (ASCs) to improve islet graft revascularization was recently reported after hybrid transplantation in mice. Within this context, we have previously shown that hyaluronan esters of butyric and retinoic acids can significantly enhance the rescuing potential of human MSCs (hMSCs). Here we evaluated whether ex vivo preconditioning of human ASCs (hASCs) with a mixture of hyaluronic (HA), butyric (BU), and retinoic (RA) acids may result in optimization of graft revascularization after islet/stem cell intrahepatic cotransplantation in syngeneic diabetic rats. We demonstrated that hASCs exposed to the mixture of molecules are able to increase the secretion of vascular endothelial growth factor (VEGF) as well as the transcription of angiogenic genes, including VEGF, KDR (kinase insert domain receptor), and hepatocyte growth factor (HGF). Rats transplanted with islets cocultured with preconditioned hASCs exhibited a better glycemic control than rats transplanted with an equal volume of islets and control hASCs. Cotransplantation with preconditioned hASCs was also associated with enhanced islet revascularization in vivo, as highlighted by graft morphological analysis. The observed increase in islet graft revascularization and function suggests that our method of stem cell preconditioning may represent a novel strategy to remarkably improve the efficacy of islets-hMSCs cotransplantation.

Urological Complications After Kidney Transplantation: Experience of More Than 1000 Transplantations

OBJECTIVE Urinary fistulas and stenoses are the most common complications that may require surgical revision after kidney transplantation. The aim of this study was to retrospectively assess the incidence of and risk factors for early (within 30 days) or late major urological complications (stenoses and fistulas) after kidney transplantation. PATIENTS AND METHODS The study population comprised 1142 consecutive patients who underwent kidney transplantation between January 1990 and September 2007. Endpoints were early and late urological complications (stenoses and fistulas). The variables considered upon multivariate and univariate analyses were: recipient age, sex, etiology of renal failure, number (first/second) and type (single/double/combined with other organs) of kidney transplantations, cold ischemia time, type of urinary reconstruction, stent positioning, as well as donor cause of death, sex, age, and serum creatinine and clearance. We also examined the presence of graft polar arteries, acute rejection episodes, and postoperative graft function. RESULTS Among 1142 transplantation performed at our center, 100 patients (8.7%) experienced 107 urological complications: 85 (79.4%) were early (56 fistulas, 29 stenoses) and 22 (20.5%) late (7 fistulas and 15 stenoses). Multivariate analysis for all complications revealed significant associations with male recipient sex (P = .00, HR = 2), while first kidney transplantation was protective (P = .00, HR = .4). Male gender both of the recipient and of the donor was significantly associated with early fistulas (P = .01, HR = 2.5 and P = .02, HR = 2, respectively). First (versus second) kidney transplantation had a protective effect on early stenoses (P = .01, HR = .27). Late fistulas were associated with anastomotic stenting (P = .03) in univariate but not multivariate analysis. Multivariate analysis for late stenoses did not demonstrate any significant association with the considered variables; however, the late stenosis cases showed significantly higher recipient and donor ages (P < .05) and a lower donor creatinine clearance (P < .05). The type of urinary anastomosis, stenting, cold ischemia time, presence of polar arterial branches, and type of transplantation did not influence the incidence of urinary fistulas or stenoses. CONCLUSIONS Our data confirmed that older recipients and organs from older donors, especially of male gender, and retransplantations are to be considered risk factors for urological complications. The present analysis cannot suggest any modification of the actual surgical strategy that would prevent urological complications in kidney transplantation.

Hepatic venous pressure gradient in the preoperative assessment of patients with resectable hepatocellular carcinoma

BACKGROUNDS & AIMS To assess the relationship existing between hepatic venous pressure gradient (HVPG) and the occurrence of post-hepatectomy liver failure (PHLF) grade B/C after resection of hepatocellular carcinoma (HCC) and persistent worsening of liver function. METHODS Data from 70 consecutive prospectively enrolled HCC patients undergoing resection were collected and analysed. PHLF grade B/C was defined by the International Study Group of Liver Surgery recommendations. The appearance of unresolved decompensation was also analysed. RESULTS Postoperative and 90-day mortality were null. The median HVPG value was 9mmHg (range: 4-18) and the median Model for End-stage Liver Disease (MELD) score was 8 (range: 6-14); 34 patients had an HVPG ⩾10mmHg (48.6%). Forty-nine patients had an uneventful (Grade A) postoperative course, including 17 with an HVPG ⩾10mmHg (24.2% of 70 patients). Grade B complications occurred in 20 patients (3 with an HVPG <10mmHg and 17 with an HVPG ⩾10mmHg; p<0.001); only one grade C complication occurred in a patient with an HVPG <10mmHg, subsequently successfully undergoing liver transplantation. Median MELD score returned to preoperative values after a transient postoperative increase, regardless of the HVPG values; after three months, it returned to the preoperative of 8 in patients with an HVPG <10mmHg and of 9 in patients with an HVPG ⩾10mmHg (p=0.077 and 0.076 at paired test, respectively). CONCLUSIONS The hepatic venous pressure gradient can be used before surgery to stratify the risk of PHLF but the proposed cut-off of 10mmHg excludes approximately one-quarter of the patients who would benefit from surgery without short to mid-term postoperative sequelae.

Berberine and its metabolites: relationship between physicochemical properties and plasma levels after administration to human subjects.

Berberine (1) is an alkaloid used widely in the treatment of several diseases. However, its physicochemical properties, pharmacokinetics, and metabolism remain unclear, and conflicting data have been reported. In this study, the main physicochemical properties of 1 and its metabolites were evaluated, including lipophilicity, solubility, pKa, and albumin binding. A sensitive HPLC-ESIMS/MS method was developed and validated to identify 1 and its main metabolites in human plasma. This method was used to quantify their levels in the plasma of healthy volunteers and hypercholesterolemic patients following a single dose and chronic administration, respectively. In both cases, berberrubine (2) was found to be the main metabolite. Surprisingly, 2 is more lipophilic than 1, which suggests that this compound tautomerizes to a highly conjugated, electroneutral quinoid structure. This was confirmed by NMR studies. These results indicate that the higher plasma concentration of 2 was a consequence of a more efficient intestinal absorption, suggesting that berberrubine is potentially more pharmacologically active than berberine.

Immunosuppression Modifications Based on an Immune Response Assay: Results of a Randomized, Controlled Trial

Background An immune function assay shows promise for identifying solid organ recipients at risk for infection or rejection. The following randomized prospective study was designed to assess the clinical benefits of adjusting immunosuppressive therapy in liver recipients based on immune function assay results. Methods Adult liver recipients were randomized to standard practice (control group; n = 102) or serial immune function testing (interventional group; n = 100) performed with a commercially available in vitro diagnostic assay (ImmuKnow; Viracor-IBT Laboratories, Lee’s Summit, MO) before transplantation, immediately after surgery and at day 1, weeks 1 to 4, 6, and 8, and months 3 to 6, 9, and 12. The assay was repeated within 7 days of suspected/confirmed rejection/infection and within 1 week after event resolution. Results Based on immune function values, tacrolimus doses were reduced 25% when values were less than 130 ng/mL adenosine triphosphate (low immune cell response) and increased 25% when values were greater than 450 ng/mL adenosine triphosphate (strong immune cell response). The 1-year patient survival was significantly higher in the interventional arm (95% vs 82%; P < 0.01) and the incidence of infections longer than 14 days after transplantation was significantly lower among patients in the interventional arm (42.0% vs. 54.9%, P < 0.05). The difference in infection rates was because of lower bacterial (32% vs 46%; P < 0.05) and fungal infection (2% vs 11%; P < 0.05). Among recipients without adverse events, the study group had lower tacrolimus dosages and blood levels. Conclusions Immune function testing provided additional data which helped optimize immunosuppression and improve patient outcomes.

Semisynthetic Bile Acid FXR and TGR5 Agonists: Physicochemical Properties, Pharmacokinetics, and Metabolism in the Rat

We report on the relationship between the structure-pharmacokinetics, metabolism, and therapeutic activity of semisynthetic bile acid analogs, including 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid (a selective farnesoid X receptor [FXR] receptor agonist), 6α-ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid (a specific Takeda G protein–coupled receptor 5 [TGR5] receptor agonist), and 6α-ethyl-3α,7α-dihydroxy-24-nor-5β-cholan-23-sulfate (a dual FXR/TGR5 agonist). We measured the main physicochemical properties of these molecules, including ionization constants, water solubility, lipophilicity, detergency, and protein binding. Biliary secretion and metabolism and plasma and hepatic concentrations were evaluated by high-pressure liquid chromatography- electrospray-mass spectrometry/mass spectrometry in bile fistula rat and compared with natural analogs chenodeoxycholic, cholic acid, and taurochenodexycholic acid and intestinal bacteria metabolism was evaluated in terms of 7α-dehydroxylase substrate-specificity in anaerobic human stool culture. The semisynthetic derivatives detergency, measured in terms of their critical micellar concentration, was quite similar to the natural analogs. They were slightly more lipophilic than the corresponding natural analogs, evaluated by their 1-octanol water partition coefficient (log P), because of the ethyl group in 6 position, which makes these molecules very stable toward bacterial 7-dehydroxylation. The hepatic metabolism and biliary secretion were different: 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid, as chenodeoxycholic acid, was efficiently conjugated with taurine in the liver and, only in this form, promptly and efficiently secreted in bile. 6α-Ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid was poorly conjugated with taurine because of the steric hindrance of the methyl at C23(S) position metabolized to the C23(R) isomer and partly conjugated with taurine. Conversely, 6α-ethyl-3α,7α-dihydroxy-24-nor-5β-cholan-23-sulfate was secreted in bile unmodified and as 3-glucuronide. Therefore, minor structural modifications profoundly influence the metabolism and biodistribution in the target organs where these analogs exert therapeutic effects by interacting with FXR and/or TGR5 receptors.

Aortoiliac surgery concomitant with kidney transplantation: a single center experience

Abstract:  Introduction:  Aortoiliac pathology in kidney allograft recipients is not rare but treatment timing is controversial. As most publications on this topic are case reports it’s difficult to evaluate long‐term outcomes of those clinical challenges. Herein we report long‐term results of these procedures.

Extracorporeal Portal Vein Oxygenation Improves Outcome of Acute Liver Failure in Swine

BACKGROUND Portal vein arterialization (PVA) has shown efficacy to treat acute liver failure (ALF) in preclinical studies. The next step is to perform large animal studies that propose a clinically acceptable method of PVA. In this study, we assessed the efficacy of PVA using an extracorporeal device to treat 2 ALF models in swine. MATERIALS AND METHODS The 2 ALF swine models were carbon tetrachloride toxic ALF and subtotal hepatectomy using 8 animals per group. PVA was performed with an extracorporeal device that may be suitable for future clinical studies. Arterial blood was drawn from the iliac artery and delivered into the portal vein for a 6-hour treatment. We analyzed biochemical, blood gas, and histological parameters as well as 1-week survival rates. RESULTS In both models, ALF was successfully achieved. Control group animals deteriorated biochemically, dropping their prothrombin times and increasing the liver enzymes. In contrast, treated animals improved with a survival rate of 75% at 7 days compared with 0% for the former group. CONCLUSIONS PVA using an extracorporeal device was feasible and effective to treat both toxic and resective ALF in swine.

Effect of Colic Vein Ligature in Rats with Loperamide-Induced Constipation

Introduction. Medical treatment in chronic constipation is not always successful. Surgery is indicated in unresponsive selected severe cases. This study presents the distal venous colic ligation in rat as a novel surgical approach. Materials and Methods. 16 rats (study group) were evaluated in 3 phases of 6 days each: A (normal conditions), B (loperamide-induced constipation), and C (colic vein legation) and compared with rats treated in phase C with PEG 4,000 (control group). Blood biochemical and physiological parameters, daily fecal water content (FWC), and histological analysis were performed in all study phases. Results. No biochemical and physiological parameters changes were observed. FWC decreased in phase B and increased in phase C in both groups with a grow up to 2.3-fold in study group compared to control (P < 0.0001). Moreover, in study group, a high number of colonic goblet cells were detected (phase C versus phase B: P < 0.001) while no differences were registered in control. Conclusion. By ligature of the colic vein in constipated rats, an increase in FWC and goblet cells higher than in PEG treated rats was detected. The described surgical procedure appeared effective, simple, and safe; further studies in animal models, however, are necessary to assess its clinical applicability.

Endolymphatic immunotherapy in inoperable hepatocellular carcinoma.

INTRODUCTION We report the preliminary results of endolymphatic immunotherapy in patients with inoperable hepatocellular carcinoma (HCC). METHODS From 2003 to 2005 we enrolled 31 patients with inoperable HCC. The patients underwent monthly endolymphatic injections of 15-30 x 10(6) interleukin-2 (IL-2)-activated peripheral autologous lymphocytes (LAK) and 250 IU of IL-2. Follow-up included blood biochemistry every 3 months and imaging studies every 6 months. To assess therapy efficacy we considered 12 biochemical parameters, vascular invasion or thrombosis, Child-Pugh scoring system, histological grading, lymphadenopathy, viral state, and alpha-fetoprotein. RESULTS Sixteen patients completed at least 3 cycles, and 10 patients completed more than 6. No clinically significant adverse reactions occurred. Imaging studies showed no significant decrease in tumor mass. However, the survival of patients who completed 12 therapy cycles was significantly higher than survival of patients with fewer than 12 cycles. Both are significantly higher than that of untreated patients. All patients with 12 completed cycles showed an improvement of 9 parameters or more. DISCUSSION Endolymphatic immunotherapy is safe, easily performed, inexpensive, and effective in terms of survival. This study should encourage future large-scale investigations so as to reach a firmer conclusion and define uniform inclusion criteria.