Systematic Sampling of the Female Reproductive System for Molecular Characterization.

As part of the National Institutes of Health Human BioMolecular Atlas Program to develop a global platform to map the 37 trillion cells in the adult human body, we are generating a comprehensive molecular characterization of the female reproductive system. Data gathered from multiple single-cell/single-nucleus and spatial molecular assays will be used to build a 3D molecular atlas. Herein, we describe our multistep protocol, beginning with an optimized organ procurement workflow that maintains functional characteristics of the uterus, ovaries, and fallopian tubes by perfusing these organs with preservation solution. We have also developed a structured tissue sampling procedure that retains information on individual-level anatomic, physiologic, and individual diversity of the female reproductive system, toward full exploration of the function and structure of female reproductive cells. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Preparation and preservation of the female reproductive system (ovaries, fallopian tubes, and uterus) prior to procurement Basic Protocol 2: Removal of the female reproductive system en bloc Basic Protocol 3: Postsurgical dissection of ovaries Basic Protocol 4: Postsurgical dissection of fallopian tubes Basic Protocol 5: Postsurgical dissection of cervix Basic Protocol 6: Postsurgical dissection of uterine body Support Protocol 1: OCT-embedded tissue protocol Support Protocol 2: Tissue fixation protocol Support Protocol 3: Snap-frozen tissue protocol Basic Protocol 7: Tissue slice preparation for Visium analysis Support Protocol 4: Hematoxylin and eosin staining for 10X Visium imaging Basic Protocol 8: Manual tissue dissociation for Multiome analysis Basic Protocol 9: Tissue dissociation for Multiome analysis using S2 Singulator.

A novel role for the Wnt inhibitor APCDD1 in adipocyte differentiation: Implications for diet-induced obesity.

Impaired adipogenic differentiation during diet-induced obesity (DIO) promotes adipocyte hypertrophy and inflammation, thereby contributing to metabolic disease. Adenomatosis polyposis coli down-regulated 1 (APCDD1) has recently been identified as an inhibitor of Wnt signaling, a key regulator of adipogenic differentiation. Here we report a novel role for APCDD1 in adipogenic differentiation via repression of Wnt signaling and an epigenetic linkage between miR-130 and APCDD1 in DIO. APCDD1 expression was significantly up-regulated in mature adipocytes compared with undifferentiated preadipocytes in both human and mouse subcutaneous adipose tissues. siRNA-based silencing of APCDD1 in 3T3-L1 preadipocytes markedly increased the expression of Wnt signaling proteins (Wnt3a, Wnt5a, Wnt10b, LRP5, and ß-catenin) and inhibited the expression of adipocyte differentiation markers (CCAAT/enhancer-binding protein α (C/EBPα) and peroxisome proliferator-activated receptor γ (PPARγ)) and lipid droplet accumulation, whereas adenovirus-mediated overexpression of APCDD1 enhanced adipogenic differentiation. Notably, DIO mice exhibited reduced APCDD1 expression and increased Wnt expression in both subcutaneous and visceral adipose tissues and impaired adipogenic differentiation in vitro Mechanistically, we found that miR-130, whose expression is up-regulated in adipose tissues of DIO mice, could directly target the 3'-untranslated region of the APCDD1 gene. Furthermore, transfection of an miR-130 inhibitor in preadipocytes enhanced, whereas an miR-130 mimic blunted, adipogenic differentiation, suggesting that miR-130 contributes to impaired adipogenic differentiation during DIO by repressing APCDD1 expression. Finally, human subcutaneous adipose tissues isolated from obese individuals exhibited reduced expression of APCDD1, C/EBPα, and PPARγ compared with those from non-obese subjects. Taken together, these novel findings suggest that APCDD1 positively regulates adipogenic differentiation and that its down-regulation by miR-130 during DIO may contribute to impaired adipogenic differentiation and obesity-related metabolic disease.

Ex Situ Perfusion of Human Limb Allografts for 24 Hours.

BACKGROUND: Vascularized composite allografts, particularly hand and forearm, have limited ischemic tolerance after procurement. In bilateral hand transplantations, this demands a 2 team approach and expedited transfer of the allograft, limiting the recovery to a small geographic area. Ex situ perfusion may be an alternative allograft preservation method to extend allograft survival time. This is a short report of 5 human limbs maintained for 24 hours with ex situ perfusion. METHODS: Upper limbs were procured from brain-dead organ donors. Following recovery, the brachial artery was cannulated and flushed with 10 000 U of heparin. The limb was then attached to a custom-made, near-normothermic (30-33°C) ex situ perfusion system composed of a pump, reservoir, and oxygenator. Perfusate was plasma-based with a hemoglobin concentration of 4 to 6 g/dL. RESULTS: Average warm ischemia time was 76 minutes. Perfusion was maintained at an average systolic pressure of 93 ± 2 mm Hg, flow 310 ± 20 mL/min, and vascular resistance 153 ± 16 mm Hg/L per minute. Average oxygen consumption was 1.1 ± 0.2 mL/kg per minute. Neuromuscular electrical stimulation continually displayed contraction until the end of perfusion, and histology showed no myocyte injury. CONCLUSIONS: Human limb allografts appeared viable after 24 hours of near-normothermic ex situ perfusion. Although these results are early and need validation with transplantation, this technology has promise for extending allograft storage times.

Echo Decorrelation Imaging of Rabbit Liver and VX2 Tumor during In Vivo Ultrasound Ablation.

In open surgical procedures, image-ablate ultrasound arrays performed thermal ablation and imaging on rabbit liver lobes with implanted VX2 tumor. Treatments included unfocused (bulk ultrasound ablation, N = 10) and focused (high-intensity focused ultrasound ablation, N = 13) exposure conditions. Echo decorrelation and integrated backscatter images were formed from pulse-echo data recorded during rest periods after each therapy pulse. Echo decorrelation images were corrected for artifacts using decorrelation measured prior to ablation. Ablation prediction performance was assessed using receiver operating characteristic curves. Results revealed significantly increased echo decorrelation and integrated backscatter in both ablated liver and ablated tumor relative to unablated tissue, with larger differences observed in liver than in tumor. For receiver operating characteristic curves computed from all ablation exposures, both echo decorrelation and integrated backscatter predicted liver and tumor ablation with statistically significant success, and echo decorrelation was significantly better as a predictor of liver ablation. These results indicate echo decorrelation imaging is a successful predictor of local thermal ablation in both normal liver and tumor tissue, with potential for real-time therapy monitoring.

HDAC9 knockout mice are protected from adipose tissue dysfunction and systemic metabolic disease during high-fat feeding.

During chronic caloric excess, adipose tissue expands primarily by enlargement of individual adipocytes, which become stressed with lipid overloading, thereby contributing to obesity-related disease. Although adipose tissue contains numerous preadipocytes, differentiation into functionally competent adipocytes is insufficient to accommodate the chronic caloric excess and prevent adipocyte overloading. We report for the first time that a chronic high-fat diet (HFD) impairs adipogenic differentiation, leading to accumulation of inefficiently differentiated adipocytes with blunted expression of adipogenic differentiation-specific genes. Preadipocytes from these mice likewise exhibit impaired adipogenic differentiation, and this phenotype persists during in vitro cell culture. HFD-induced impaired adipogenic differentiation is associated with elevated expression of histone deacetylase 9 (HDAC9), an endogenous negative regulator of adipogenic differentiation. Genetic ablation of HDAC9 improves adipogenic differentiation and systemic metabolic state during an HFD, resulting in diminished weight gain, improved glucose tolerance and insulin sensitivity, and reduced hepatosteatosis. Moreover, compared with wild-type mice, HDAC9 knockout mice exhibit upregulated expression of beige adipocyte marker genes, particularly during an HFD, in association with increased energy expenditure and adaptive thermogenesis. These results suggest that targeting HDAC9 may be an effective strategy for combating obesity-related metabolic disease.

In vivo thermal ablation monitoring using ultrasound echo decorrelation imaging.

Previous work indicated that ultrasound echo decorrelation imaging can track and quantify changes in echo signals to predict thermal damage during in vitro radiofrequency ablation (RFA). In the in vivo studies reported here, the feasibility of using echo decorrelation imaging as a treatment monitoring tool was assessed. RFA was performed on normal swine liver (N = 5), and ultrasound ablation using image-ablate arrays was performed on rabbit liver implanted with VX2 tumors (N = 2). Echo decorrelation and integrated backscatter were computed from Hilbert transformed pulse-echo data acquired during RFA and ultrasound ablation treatments. Receiver operating characteristic (ROC) curves were employed to assess the ability of echo decorrelation imaging and integrated backscatter to predict ablation. Area under the ROC curves (AUROC) was determined for RFA and ultrasound ablation using echo decorrelation imaging. Ablation was predicted more accurately using echo decorrelation imaging (AUROC = 0.832 and 0.776 for RFA and ultrasound ablation, respectively) than using integrated backscatter (AUROC = 0.734 and 0.494).

Human coronary artery perivascular adipocytes overexpress genes responsible for regulating vascular morphology, inflammation, and hemostasis.

Inflammatory cross talk between perivascular adipose tissue and the blood vessel wall has been proposed to contribute to the pathogenesis of atherosclerosis. We previously reported that human perivascular (PV) adipocytes exhibit a proinflammatory phenotype and less adipogenic differentiation than do subcutaneous (SQ) adipocytes. To gain a global view of the genomic basis of biologic differences between PV and SQ adipocytes, we performed genome-wide expression analyses to identify differentially expressed genes between adipocytes derived from human SQ vs. PV adipose tissues. Although >90% of well-expressed genes were similarly regulated, we identified a signature of 307 differentially expressed genes that were highly enriched for functions associated with the regulation of angiogenesis, vascular morphology, inflammation, and blood clotting. Of the 156 PV upregulated genes, 59 associate with angiogenesis, vascular biology, or inflammation, noteworthy of which include TNFRSF11B (osteoprotegerin), PLAT, TGFB1, THBS2, HIF1A, GATA6, and SERPINE1. Of 166 PV downregulated genes, 21 associated with vascular biology and inflammation, including ANGPT1, ANGPTL1, and VEGFC. Consistent with the emergent hypothesis that PV adipocytes differentially regulate angiogenesis and inflammation, cell culture-derived adipocyte-conditioned media from PV adipocytes strongly enhanced endothelial cell tubulogenesis and monocyte migration compared with media from SQ adipocytes. These findings demonstrate that PV adipocytes have the potential to significantly modulate vascular inflammatory crosstalk in the setting of atherosclerosis by their ability to signal to both endothelial and inflammatory cells.

Allele-specific expression of angiotensinogen in human subcutaneous adipose tissue.

The angiotensinogen gene is genetically linked with hypertension, but the mechanistic basis for association of sequence variants in the promoter and coding region of the gene remains unclear. An E-box at position -20 has been hypothesized to control the level of angiotensinogen expression, but its mechanistic importance for angiotensinogen expression in human tissues is uncertain. We developed an allele-specific polymerase chain reaction-based assay to distinguish between angiotensinogen mRNA derived from variants at the -20 position (rs5050) in the angiotensinogen promoter in adipose tissues obtained during surgery. The assay takes advantage of linkage disequilibrium between the rs5050 (located in the promoter) and rs4762 (located in the coding region) single nucleotide polymorphisms. This strategy allowed us to assess the level of allele-specific expression in A-20C heterozygous subjects comparing the relative proportion of each allele with the total, thus eliminating the problem of variability in the level of total angiotensinogen mRNA among subjects. We show that angiotensinogen mRNA derived from the -20C allele is expressed significantly higher than that derived from the -20A allele in subcutaneous adipose tissue, and increased expression correlates with enriched chromatin binding of upstream stimulatory factor-2 to the -20C E-box compared with -20A. This may be depot selective because we were unable to detect these differences in omental adipose. This provides the first data directly comparing expression of angiotensinogen mRNA and differential transcription factor binding derived from 2 variant alleles in human tissue where the ratio of expression of one allele to another can be accurately determined.

Laparoscopic cholecystectomy in the cirrhotic patient: predictors of outcome.

BACKGROUND: We sought to determine the outcome predictors of 94 cirrhotic patients undergoing laparoscopic cholecystectomy (LC). METHODS: We performed a single-center, retrospective review of cirrhotic patients undergoing LC for symptomatic gallbladder disease. Statistical analysis was completed using the Chi-square, Wilcoxon rank-sum, and Student t tests as appropriate. RESULTS: Ninety-four procedures were completed. The median Child-Turcotte-Pugh (CTP) score was 6 (range, 5-12), and the average Model for End-Stage Liver Disease (MELD) score was 11 ± 5. Hepatitis C was the most common etiology of liver disease (50%) followed by Laennec's cirrhosis (22%). The average length of stay was 2.6 ± 4.3 days; 21% were outpatient procedures. The conversion rate was 11%. Conversion risk factors were decreased serum albumin, increased MELD score, and blood loss. Morbidity occurred in 32 patients. Predictors of morbidity were decreases in serum albumin, increases in International Normalized Ratio (INR) and CTP score, and the number of intraoperative red blood cell transfusions. Mortality occurred in 4 patients. Increased INR, CTP score, CTP class, the number of intraoperative blood and platelet transfusions were predictors of mortality. CONCLUSION: LC can be safely performed in cirrhotic patients with appropriate patient selection. Liver synthetic function, operative blood loss, transfusion requirement, CTP, and MELD scores may be used to predict outcomes in these patients.

Results of live donor liver transplantation in patients with hepatitic C virus infection: the HCV 3 trial experience.

Chronic hepatitis C virus (HCV) is the most common disease indication for liver transplantation (LT). Outcomes are compromised by near universal recurrence of HCV. A prospective multi-center randomized study to evaluate immunosuppressive strategies in HCV+ transplant recipients provided the opportunity to assess impact of live donor (LD) LT. Two hundred and ninety-five patients undergoing LT for HCV (260 deceased donor [DD] recipients/35 LD recipients), randomized to three regimens, were followed for two yr for patient and graft survival and rate and severity of recurrent HCV. Biopsies were performed at baseline, 3, 12, and 24 months. One- and two-yr patient survival for LD recipients was 88.1% and 81.1% vs. 90.5% and 84.6% for DD recipients (p = 0.5665). One- and two-yr graft survival for LD recipients was 82.9% and 76.2% vs. 87.9% and 81.7% for DD recipients (p = 0.3921). Recurrent HCV did not account for more deaths or graft losses in the LD recipients. In this prospective study, controlled for immunosuppression, use of LD organs did not increase the rate or severity of HCV recurrence. The more elective nature of LDLT affords an opportunity to manipulate donor and recipient factors that can impact upon outcomes.

Treatment of rabbit liver cancer in vivo using miniaturized image-ablate ultrasound arrays.

In the preclinical studies reported here, VX2 cancer within rabbit liver has been treated by bulk ultrasound ablation employing miniaturized image-ablate arrays. Array probes were constructed with 32 elements in a 2.3 × 20 mm(2) aperture, packaged within a 3.1 mm stainless steel tube with a cooling and coupling balloon for in vivo use. The probes were measured capable of 50% fractional bandwidth for pulse-echo imaging (center frequency 4.4 MHz) with >110 W/cm(2) surface intensity available at sonication frequencies 3.5 and 4.8 MHz. B-scan imaging performance of the arrays was measured to be comparable to larger diagnostic linear arrays, although nearfield image quality was reduced by ringdown artifacts. A series of in vivo ablation procedures was performed using an unfocused 32-element aperture firing at 4.8 MHz with exposure durations 20-70.5 s and in situ spatial average, temporal average intensities 22.4-38.5 W/cm(2). Ablation of a complete tumor cross-section was confirmed by vital staining in seven of 12 exposures, with four exposures ablating an additional margin >1 mm beyond the tumor in all directions. Analysis suggests a threshold ablation effect, with complete ablation of tumor cross-sections for exposures with delivery of >838 J acoustic energy. The results show feasibility for in vivo liver cancer ablation using miniaturized image-ablate arrays suitable for interstitial deployment.

Histone deacetylase 9 is a negative regulator of adipogenic differentiation.

Differentiation of preadipocytes into mature adipocytes capable of efficiently storing lipids is an important regulatory mechanism in obesity. Here, we examined the involvement of histone deacetylases (HDACs) and histone acetyltransferases (HATs) in the regulation of adipogenesis. We find that among the various members of the HDAC and HAT families, only HDAC9 exhibited dramatic down-regulation preceding adipogenic differentiation. Preadipocytes from HDAC9 gene knock-out mice exhibited accelerated adipogenic differentiation, whereas HDAC9 overexpression in 3T3-L1 preadipocytes suppressed adipogenic differentiation, demonstrating its direct role as a negative regulator of adipogenesis. HDAC9 expression was higher in visceral as compared with subcutaneous preadipocytes, negatively correlating with their potential to undergo adipogenic differentiation in vitro. HDAC9 localized in the nucleus, and its negative regulation of adipogenesis segregates with the N-terminal nuclear targeting domain, whereas the C-terminal deacetylase domain is dispensable for this function. HDAC9 co-precipitates with USF1 and is recruited with USF1 at the E-box region of the C/EBPα gene promoter in preadipocytes. Upon induction of adipogenic differentiation, HDAC9 is down-regulated, leading to its dissociation from the USF1 complex, whereas p300 HAT is up-regulated to allow its association with USF1 and accumulation at the E-box site of the C/EBPα promoter in differentiated adipocytes. This reciprocal regulation of HDAC9 and p300 HAT in the USF1 complex is associated with increased C/EBPα expression, a master regulator of adipogenic differentiation. These findings provide new insights into mechanisms of adipogenic differentiation and document a critical regulatory role for HDAC9 in adipogenic differentiation through a deacetylase-independent mechanism.

The impact of a change in tacrolimus monitoring immunoassay techniques on clinical decision making.

CONTEXT: Tacrolimus is an immunosuppressant that undergoes therapeutic drug monitoring. The laboratory at our institution changed its immunoassay techniques from the fluorescence polarization immunoassay to the cloned enzyme donation immunoassay. OBJECTIVE: To evaluate the relationship between the 2 assays and to determine the impact of the change on clinical decision making. DESIGN: A retrospective study of patients admitted to the hospital during the assay transition period. Tacrolimus values for the 2 assays were collected for 4 weeks and compared. SETTING: An academic health center. PATIENTS: Liver transplant patients hospitalized from February 18, 2008, to March 18, 2008. MAIN OUTCOME MEASURE: The primary outcome was the agreement between the results of the 2 immunoassays. Secondary outcome was agreement of clinical decision making with established patient-specific therapeutic ranges or with a 30% difference in absolute values between the assays. RESULTS: Seventy-nine pairs of tacrolimus concentrations were collected from 21 liver transplant patients. The mean (SD) tacrolimus concentrations were 7.36 (4.21) microg/L for the fluorescence polarization immunoassay and 9.00 (5.30) microg/L for the cloned enzyme donation immunoassay (P = .03). A clinically different decision would have been made if the fluorescence polarization immunoassay value had not been reported 51% of the time. A Bland-Altman plot indicated no relationship between the assay results. CONCLUSION: A change in tacrolimus monitoring assay would have resulted in different clinical decisions 51% of the time. Awareness of changes in assay technology must be heightened to enhance clinical decision making and prevent potential impact on morbidity among liver transplant patients.

Proinflammatory phenotype of perivascular adipocytes: influence of high-fat feeding.

Adipose tissue depots originate from distinct precursor cells, are functionally diverse, and modulate disease processes in a depot-specific manner. However, the functional properties of perivascular adipocytes, and their influence on disease of the blood vessel wall, remain to be determined. We show that human coronary perivascular adipocytes exhibit a reduced state of adipocytic differentiation as compared with adipocytes derived from subcutaneous and visceral (perirenal) adipose depots. Secretion of antiinflammatory adiponectin is markedly reduced, whereas that of proinflammatory cytokines interleukin-6, interleukin-8, and monocyte chemoattractant protein-1, is markedly increased in perivascular adipocytes. These depot-specific differences in adipocyte function are demonstrable in both freshly isolated adipose tissues and in vitro-differentiated adipocytes. Murine aortic arch perivascular adipose tissues likewise express lower levels of adipocyte-associated genes as compared with subcutaneous and visceral adipose tissues. Moreover, 2 weeks of high-fat feeding caused further reductions in adipocyte-associated gene expression, while upregulating proinflammatory gene expression, in perivascular adipose tissues. These changes were observed in the absence of macrophage recruitment to the perivascular adipose depot. We conclude that perivascular adipocytes exhibit reduced differentiation and a heightened proinflammatory state, properties that are intrinsic to the adipocytes residing in this depot. Dysfunction of perivascular adipose tissue induced by fat feeding suggests that this unique adipose depot is capable of linking metabolic signals to inflammation in the blood vessel wall.

Ultrasound monitoring of in vitro radio frequency ablation by echo decorrelation imaging.

OBJECTIVE: The purpose of this study was to test ultrasound echo decorrelation imaging for mapping and characterization of tissue effects caused by radio frequency ablation (RFA). METHODS: Radio frequency ablation procedures (6-minute duration, 20-W power) were performed on fresh ex vivo bovine liver tissue (n = 9) with continuous acquisition of beam-formed ultrasound echo data from a 7-MHz linear array. Echo data were processed to form B-scan images, echo decorrelation images (related to rapid random changes in echo waveforms), and integrated backscatter images (related to local changes in received echo energy). Echo decorrelation and integrated backscatter values at the location of a low-noise thermocouple were assessed as functions of temperature. Echo decorrelation and integrated backscatter images were directly compared with ablated tissue cross sections and quantitatively evaluated as predictors of tissue ablation and overtreatment. RESULTS: Echo decorrelation maps corresponded with local tissue temperature and ablation effects. Consistent echo decorrelation increases were observed for temperatures above 75 degrees C, whereas integrated backscatter maps showed a nonmonotonic temperature dependence complicated by acoustic shadowing, with high variance at large temperature elevations. In receiver operating characteristic curve analysis of echo decorrelation and integrated backscatter maps as predictors of local tissue ablation, echo decorrelation performed well (area under the receiver operating characteristic curve [AUROC] = 0.855 for ablation and 0.913 for overtreatment), whereas integrated backscatter performed poorly (AUROC < 0.6). CONCLUSIONS: Echo decorrelation imaging can map tissue changes due to RFA in vitro, with local echo decorrelation corresponding strongly to local tissue temperature elevations and ablation effects. With further development and in vivo validation, echo decorrelation imaging is potentially useful for improved image guidance of clinical RFA procedures.

Experience with more than 500 minimally invasive hepatic procedures.

OBJECTIVE: To evaluate our experience with more than 500 minimally invasive hepatic procedures. SUMMARY BACKGROUND DATA: Recent data have confirmed the safety and efficacy of minimally invasive liver surgery. Despite these reports, no programmatic approach to minimally invasive liver surgery has been proposed. METHODS: We retrospectively reviewed all patients who underwent a minimally invasive procedure for the management of hepatic tumors between January 2001 and April 2008. Patients were divided into 3 groups: laparoscopy with intraoperative ultrasound and biopsy only, laparoscopic radiofrequency ablation (RFA), and minimally invasive resection. To compare the various forms of surgery, we analyzed the incidence of complications, tumor recurrence, mortality, and cost. Statistical analysis was performed using chi(2) analysis, Student t test, Kaplan-Meier survival analysis with the log-rank test, and multivariable Cox models. RESULTS: A total of 590 minimally invasive hepatic procedures were performed during 489 operative interventions. The representative tumor histologies were: hepatocellular carcinoma (HCC; N = 210), colorectal carcinoma (N = 40), miscellaneous liver metastases (N = 42), biliary cancer (N = 20), and benign tumors (N = 176). Thirty-five patients underwent laparoscopic ultrasound and confirmatory biopsy alone; 201 patients underwent 240 laparoscopic RFAs, and 253 patients underwent 306 minimally invasive resections. Conversion rates to open surgery for the RFA and resection group were 2% overall. One hundred ninety-nine (40.6%) patients were cirrhotic; 31 resections were performed in cirrhotic patients. Complication and mortality rates for RFA and resection were comparable (11% vs. 16%, and 1.5% vs. 1.6%). However, complication rates (14% vs. 29%; P = 0.02) and mortality (0.3% vs. 9.7%; P = 0.006) rates were higher in the cirrhotic versus noncirrhotic resection group. Overall recurrence rates for RFA and resection groups were 24% and 23%, respectively. Local recurrence rates were higher in the RFA group (6.3% versus 1.5%; P < 0.06). Overall patient survival differed between HCC patients receiving RFA alone and those receiving RFA and OLT (P < 0.0001). Overall survival for cancer patients receiving RFA versus resection differed significantly when unadjusted for other covariates (P = 0.01), and remained marginally significant in a multivariable model (P = 0.056). CONCLUSIONS: Minimally invasive hepatic surgery has become a viable alternative to open hepatic surgery. Our present data are equivalent or superior to those encountered in any large open series. Our experience with RFA confirms a low local recurrence rate and an excellent technique for bridging patients to transplantation. Morbidity and mortality rates for minimally invasive hepatic resections in cirrhotics, is similar to other reported open resection series. This series confirmed excellent interim survival rates after laparoscopic HR and superiority over RFA in the treatment of cancer, with significantly lower local tumor recurrence rate.

Incidence of adverse events with HMG-CoA reductase inhibitors in liver transplant patients.

UNLABELLED: Transplant patients are at increased risk of developing dyslipidemia, which contributes to coronary artery disease and cardiovascular events. The purpose of this study was to explore documented adverse effects of liver transplant recipients receiving lipid-lowering therapies. METHODS: A retrospective chart review of 69 liver transplant patients was conducted to evaluate the incidence of adverse effects, especially rhabdomyolysis and liver function abnormalities, in liver transplant patients treated with a lipid lowering agent (LLA). Data were collected from the time of initiation of LLA to 12 months later, looking at the type, dose, and duration of LLA, concurrent cytochrome P450 inhibitors, immunosuppression used, and laboratory parameters. RESULTS: For HMG-CoA reductase inhibitor therapy, simvistatin was used in five (7.8%) patients, pravastatin in 40 (62.5%), fluvastatin in one (1.6%), atorvastatin in five (7.8%), and lovastatin in three (4.7%). Gemfibrozil, a fibric acid derivative, was employed as monotherapy in 10 (15.6%) of patients. There were five patients who received combination therapy with a fibric acid derivative, four (80%) with gemfibrozil + pravastatin, and one (20%) with gemfibrozil + simvastatin. Six patients studied had adverse effects, five (7.2%) with myalgia and one (1.4%) with myopathy. LLA monotherapy with either pravastatin or atorvastatin was used in these patients. The five patients with myalgia were on concurrent therapy with cyclosporin, and the patient with myopathy was on concurrent cyclosporin + diltiazem therapy, both of which are P450 inhibitors. One out of 23 patients on a non-immunosuppressant P450 inhibitor developed adverse effects. No significant elevation of alanine aminotransferase, aspartate aminotransferase, or alkaline phosphatase was noted in any patient. CONCLUSIONS: Overall, there was a general tolerability with a low incidence of adverse events, no incidence of severe complications, and no alterations in liver function tests in the study population with the use of LLA.

Venous stenosis in a pig arteriovenous fistula model--anatomy, mechanisms and cellular phenotypes.

BACKGROUND: Haemodialysis vascular access dysfunction is currently a huge clinical problem. Although arteriovenous (AV) fistulae are the preferred mode of dialysis access, they have significant problems with both early (failure to mature) and late fistula failure. Both are characterized radiologically as a stenosis of the venous segment. Despite the magnitude of the clinical problem, the exact pathogenesis of AV fistula failure remains unclear. The aim of this study was to develop and validate a pig model of AV fistula stenosis and then use it to dissect out the mechanisms responsible for this lesion. METHODS: AV fistulae were created between the femoral artery and vein of Yorkshire Cross pigs. Animals were sacrificed at 2 days, 7 days, 28 days and 42 days post-surgery. At the time of sacrifice the entire specimen was divided into four regions; the arterial (AV-A) and venous (AV-V) portions of the AV anastomosis, the juxta-anastomotic segment (JA) and the proximal vein (PV), and assessed for the degree of intima-media thickening and the presence of specific cellular phenotypes. Haemodynamic parameters were not measured in this set of experiments. RESULTS: Significant luminal stenosis and intima-media thickening were present as early as 28 days and 42 days post-surgery in the pig model. In addition, within specimens from a single time point, these two parameters were maximal within the proximal vein and juxta-anastomotic segment as compared to the AV anastomosis (P < 0.0001). The vast majority of cells within the region of intima-media thickening were myofibroblasts. CONCLUSIONS: These studies suggest that early and aggressive intima-media thickening (which is made up primarily of myofibroblasts) plays an important role in AV fistula stenosis in a pig model of AV fistula placement. Interventions that target the mechanisms and cellular phenotypes described in this model, may be effective in reducing the very significant morbidity and economic costs currently associated with AV fistula failure.

Corticosteroid-free immunosuppression with daclizumab in HCV(+) liver transplant recipients: 1-year interim results of the HCV-3 study.

This work is a 1-yr interim analysis of a prospective, randomized, multicenter trial evaluating the effect of corticosteroid-free immunosuppression on hepatitis C virus-positive (HCV(+)) liver transplant recipients following liver transplantation (LT). Patients received tacrolimus and corticosteroids (Arm 1; n = 80); tacrolimus, corticosteroids, and mycophenolate mofetil (MMF) (Arm 2; n = 79); or daclizumab induction, tacrolimus, and MMF (Arm 3; n = 153). At 1 yr, 64.1%, 63.4%, and 69.4% of patients achieved the composite primary endpoint of freedom from rejection, freedom from HCV recurrence, and freedom from treatment failure, respectively. Excellent patient and graft survival did not differ significantly among treatment arms. Freedom from HCV recurrence at 1 yr was 61.8 +/- 6.2%, 60.1 +/- 6.1%, and 67.0 +/- 4.3% in Arms 1, 2, and 3, respectively (P = not significant). Freedom from rejection was significantly higher in Arm 3 compared to Arm 1 (93.0 +/- 2.2% vs. 81.9 +/- 4.4%; P = 0.011). Multivariate analysis identified acute rejection (hazard ratio = 2.692; P = 0.001) and donor age (hazard ratio = 1.015; P = 0.001) as significant risk factors for HCV recurrence. HCV recurrence was not influenced by recipient demographics, HCV genotype, or immunosuppression. In conclusion, these results suggest that a corticosteroid-free regimen of tacrolimus and MMF following daclizumab induction is safe and effective in HCV(+) liver transplant recipients.