Hepatic arterial variations and liver-related diseases of 100 consecutive donors

We prospectively studied anatomical variations and diseases of the liver in 100 consecutive donor operations during a period of 1 year. The normal arterial anatomy with a single hepatic artery (HA) from the celiac trunk was seen in 76% of all cases. Seven of twelve different major variations of the HA may be considered as rare, one of which cannot be found in the earlier literature. During harvesting, 6% of the livers were discarded, 3% on the basis of infection and 1% because of a polycystic disease. Two cases were rejected as the liver was found to be severely hypoperfused or hypoxic in an otherwise stable donor. Severe steatosis was macroscopically and histologically diagnosed in 3% of the cases, and in three donors a benign tumour was found in the liver or in the gall bladder. Two primarily nonfunctioning livers in the present series of 94 recipient operations were retrieved from this group of severely steatotic livers. As the donor liver was totally normal in only 2 out of 3 of the cases, the present study underlines the importance of searching for extremely variable anomalies of the HA and for liver-related diseases during organ harvesting.     

Tissue microarray: an effective high-throughput method to study the placenta for clinical and research purposes.

OBJECTIVE: Tissue microarray (TMA) technology allows simultaneous examination of the expression of many molecular markers (protein, mRNA, DNA, etc.) with high-throughput. The application of this technology, to date, has been largely confined to the study of cancer. Placental pathology poses unique challenges because of the size of the organ, its complex anatomy, as well as its histological heterogeneity. The objective of this study was to assess the feasibility and efficiency of TMAs for immunohistochemistry and in situ hybridization of placental tissues. STUDY DESIGN: TMAs were constructed using an automated tissue arrayer. Standard 0.6-mm or 1-mm microarray needles were used. Villous parenchyma, basal plate, and chorioamniotic membranes were targeted in each block. Five mum-thick TMA sections underwent immunohistochemical analysis of both cytoplasmic and nuclear antigens using a panel of antibodies against a variety of cytoplasmic [cytokeratin-7, vascular endothelial growth factor (VEGF), and protein Z], membranous (endoglin), and nuclear (c-fos and c-jun) antigens. mRNA in situ hybridization for surfactant protein A (SP-A) and chromogenic in situ hybridization for the Y chromosome (DYZ1) were also performed. RESULTS: Validation of TMA immunoreactivity demonstrated comparable results with corresponding whole sections. When a two-tiered scoring system (positive/negative) was employed, there was agreement between two and three cores and whole tissue sections (kappa>0.7). When a three-tiered scoring system (negative, weak-positive, or strong-positive) was used, the data from three cores showed the highest agreement with whole tissue sections (kappa >0.7). In situ hybridization experiments for mRNA and DNA were also successful in that the signals were readily detectable. Successful transfer from the donor block to the recipient block differed according to the anatomical compartment. The transfer efficiency of villous parenchyma, basal plate, and chorioamniotic membranes were 96.9% (875/903), 76.7% (115/150), and 75.4% (224/297), respectively. CONCLUSION: TMA is a practical and effective tool for high-throughput molecular analysis of the human placenta. Duplicate and triplicate cores offer agreement with whole tissue sections for two-category distinction immunostaining. TMA also affords relevant results from in situ hybridization experiments for mRNA and DNA. The major advantages are the conservation of tissues and reagents, simultaneous comparison of molecular markers in different anatomical compartments of the placenta, and reduction of experimental error.     

Genetic and environmental control of host-gut microbiota interactions

Genetics provides a potentially powerful approach to dissect host-gut microbiota interactions. Toward this end, we profiled gut microbiota using 16s rRNA gene sequencing in a panel of 110 diverse inbred strains of mice. This panel has previously been studied for a wide range of metabolic traits and can be used for high-resolution association mapping. Using a SNP-based approach with a linear mixed model, we estimated the heritability of microbiota composition. We conclude that, in a controlled environment, the genetic background accounts for a substantial fraction of abundance of most common microbiota. The mice were previously studied for response to a high-fat, high-sucrose diet, and we hypothesized that the dietary response was determined in part by gut microbiota composition. We tested this using a cross-fostering strategy in which a strain showing a modest response, SWR, was seeded with microbiota from a strain showing a strong response, A×B19. Consistent with a role of microbiota in dietary response, the cross-fostered SWR pups exhibited a significantly increased response in weight gain. To examine specific microbiota contributing to the response, we identified various genera whose abundance correlated with dietary response. Among these, we chose Akkermansia muciniphila, a common anaerobe previously associated with metabolic effects. When administered to strain A×B19 by gavage, the dietary response was significantly blunted for obesity, plasma lipids, and insulin resistance. In an effort to further understand host-microbiota interactions, we mapped loci controlling microbiota composition and prioritized candidate genes. Our publicly available data provide a resource for future studies.Studies carried out over the last decade have revealed that gut microbiota contribute to a variety of common disorders, including obesity and diabetes (Musso et al. 2011), colitis (Devkota et al. 2012), atherosclerosis (Wang et al. 2011), rheumatoid arthritis (Vaahtovuo et al. 2008), and cancer (Yoshimoto et al. 2013). The evidence for metabolic interactions is particularly strong, as a large body of data now supports the conclusion that gut microbiota influence the energy harvest from dietary components, particularly complex carbohydrates, and that metabolites such as the short-chain fatty acids produced by gut bacteria can perturb metabolic traits, including adiposity and insulin resistance (Turnbaugh et al. 2006, 2009; Backhed et al. 2007; Wen et al. 2008; Ridaura et al. 2013). Gut microbiota communities are assembled each generation, influenced by maternal seeding, environmental factors, host genetics, and age, resulting in substantial variations in composition among individuals in human populations (Eckburg et al. 2005; Costello et al. 2009; Human Microbiome Project Consortium 2012; Goodrich et al. 2014). Most experimental studies of host-gut microbiota interactions have employed large perturbations, such as comparisons of germ-free versus conventional mice, and the significance of common variations in gut microbiota composition for disease susceptibility is still poorly understood. Furthermore, while studies with germ-free mice have clearly implicated microbiota in clinically relevant traits, it has proven difficult to identify the responsible taxa of bacteria.We now report a population-based analysis of host-gut microbiota interactions in the mouse. One of the issues we explore is the role of host genetics. Although some evidence is consistent with significant heritability of gut microbiota composition, the extent to which the host controls microbiota composition under controlled environmental conditions is unclear. We also examined the role of common variations in gut microbiota in metabolic traits such as obesity and insulin resistance and mapped loci contributing to the abundance of certain microbiota. We performed our study using a resource termed the Hybrid Mouse Diversity Panel (HMDP), consisting of about 100 inbred strains of mice that have been either sequenced or subjected to high-density genotyping (Bennett et al. 2010). The resource has several advantages for genetic analysis as compared to traditional genetic crosses. First, it allows high-resolution mapping by association rather than linkage analysis, and it has now been used for the identification of a number of novel genes underlying complex traits (Farber et al. 2011; Lavinsky et al. 2015; Parks et al. 2015; Rau et al. 2015). Second, since the strains are permanent, the data from separate studies can be integrated, allowing the development of large, publicly available databases of physiological and molecular traits relevant to a variety of clinical disorders (systems.genetics.ucla.edu and phenome.jax.org). Third, the panel is ideal for examining gene-by-environment interactions, since it is possible to examine individuals of a particular genotype under a variety of conditions (Orozco et al. 2012; Parks et al. 2013).     

Bile duct confluence: anatomic variations and its classification

Accurate knowledge of the anatomy of the bile ducts is critical for successfully hepato-biliary surgery. We describe the anatomical variations of the confluence of the bile ducts, their branches patterns, frequency and classification. From 1996 to 2011, we have collected data of the bile duct confluence. 2,032 and 1,014 anatomical variations of right and left bile ducts, respectively, were reviewed and classified according to the branching pattern. The frequencies of each type of the right hepatic duct (RHD) were as follows: Type A1—1,247 (61.3 %); Type A2—296 (14.5 %); Type A3—272 (13.3 %); Type A4—124 (6.1 %); Type A5—21 (1 %) and others—72 (3.5 %) and, for the left hepatic duct (LHD) was as follows: Type B1—773 (76.2 %); Type B2—153 (15 %); Type B3—38 (3.7 %); Type B4—9 (0.8 %); Type B5—29 (2.8 %) and others—12 (1.1 %). Atypical branching patterns of both the right and left hepatic ducts were found in 14 and 8 %, respectively. The two most common variations of the RHD were right anterior and posterior hepatic ducts join together to form the RHD and trifurcation where the RHD is absent and right anterior and posterior hepatic ducts join directly to the confluence with the LHD to form the common hepatic duct. The two most common variations in the LHD were segment IV drainage to the left and right hepatic ducts.     

Effect of Experimental Hypertriglyceridaemia on Tissue and Serum Lipoprotein Lipase Activity

Abstract Hypertriglyceridaemia was produced in rats by the intravenous infusion of Intralipid emulsion or of very low density (d < 1.006) rabbit or human lipoproteins (VLDL). Lipoprotein lipase activity was assayed, in tissues removed at the end of infusion, on serum-activated mono- and triolein emulsions at pH 8.6. Hypertriglyceridaemia resulted in a marked decrease in epididymal adipose tissue lipoprotein lipase activity and in an increase in heart enzyme activity. These changes were evident with both mono- and triolein substrates. The effects on adipose tissue enzyme activity seemed roughly dependent on the triglyceride (TG) level and, relative to TG elevation, were most pronounced in the case of VLDL infusion. Serum lipoprotein lipase activity, measured in the absence of heparin, was considerably increased suggesting that the TG-rich material "leached" the adipose tissue enzyme into the circulation. Leaching of lipoprotein lipase from adipose tissue by Intralipid emulsion or VLDL was also demonstrated in an in vitro system devoid of heparin. Contact with the TG-poor, 1.006 < d < 1.063, lipoprotein induced only a small loss in adipose tissue lipoprotein lipase activity, either in vitro or in vivo.
Intracellular lipolytic activity toward mono- and triolein, measured in adipose tissue and heart homogenates at pH 7.2 in the absence of serum, was not significantly affected by TG elevation. Thus, the observed changes in lipoprotein lipase activity seem unrelated to the intracellular lipolytic activity.
It is suggested that the low adipose tissue lipoprotein lipase activity and the retarded TG removal observed in certain hypertriglyceridaemic conditions may be secondary to the increased supply of TG-rich lipoproteins.     

Human beta-defensin-2: a natural antimicrobial peptide present in amniotic fluid participates in the host response to microbial invasion of the amniotic cavity.

OBJECTIVE: Human beta-defensin-2 (HBD-2) is a potent antimicrobial peptide that is part of the innate immune response. The purpose of this study was to determine whether HBD-2 is present in amniotic fluid and if its concentration changes with microbial invasion of the amniotic cavity (MIAC) and labor. STUDY DESIGN: Amniotic fluid was retrieved by amniocentesis from 318 patients in the following groups: (1) mid-trimester (n=75); (2) term not in labor (n=28) and in labor (n=51); (3) preterm labor and intact membranes without MIAC who delivered at term (n=36), who delivered preterm without MIAC (n=52), and preterm labor with MIAC who delivered preterm (n=25); and (4) preterm premature rupture of membranes (preterm PROM) with (n=25) and without MIAC (n=26). MIAC was defined as a positive amniotic fluid culture for microorganisms. Amniotic fluid HBD-2 concentrations were determined using a sensitive and specific ELISA. Non-parametric statistics were used for analysis. RESULTS: (1) HBD-2 was detected in all amniotic fluid samples; (2) the concentration of HBD-2 did not change with gestational age from mid-trimester to term (p=0.8); (3) intra-amniotic infection was associated with a significant increase in amniotic fluid concentrations of HBD-2 in both women with preterm labor and intact membranes, and women with preterm PROM (p<0.05 for each comparison); (4) patients with preterm labor and a negative amniotic fluid culture who delivered preterm had a higher median amniotic fluid HBD-2 concentration than those with preterm labor who delivered at term (p=0.001); and (5) among patients with preterm labor without MIAC, those who had intra-amniotic inflammation (amniotic fluid white blood cell count>100 cells per mL) had a higher median amniotic fluid concentration of HBD-2 than those without this condition (p<0.002). CONCLUSION: (1) Amniotic fluid contains HBD-2, a natural antimicrobial peptide, and this may account for some of the antimicrobial activity of amniotic fluid; (2) amniotic fluid HBD-2 concentrations are increased in women with MIAC, regardless of the membrane status (intact membranes or PROM); and (3) we propose that amniotic fluid HBD-2 is part of the innate immune system within the amniotic cavity.