Long Term Liver Engraftment of Functional Hepatocytes Obtained from Germline Cell-Derived Pluripotent Stem Cells

One of the major hurdles in liver gene and cell therapy is availability of ex vivo-expanded hepatocytes. Pluripotent stem cells are an attractive alternative. Here, we show that hepatocyte precursors can be isolated from male germline cell-derived pluripotent stem cells (GPSCs) using the hepatoblast marker, Liv2, and induced to differentiate into hepatocytes in vitro. These cells expressed hepatic-specific genes and were functional as demonstrated by their ability to secrete albumin and produce urea. When transplanted in the liver parenchyma of partially hepatectomised mice, Liv2-sorted cells showed regional and heterogeneous engraftment in the injected lobe. Moreover, approximately 50% of Y chromosome-positive, GPSC-derived cells were found in the female livers, in the region of engraftment, even one month after cell injection. This is the first study showing that Liv2-sorted GPSCs-derived hepatocytes can undergo long lasting engraftment in the mouse liver. Thus, GPSCs might offer promise for regenerative medicine.     

Melanin content and MC1R function independently affect UVR-induced DNA damage in cultured human melanocytes

Malignant transformation of melanocytes leads to melanoma, the most fatal form of skin cancer. Ultraviolet radiation (UVR)-induced DNA photoproducts play an important role in melanomagenesis. Cutaneous melanin content represents a major photoprotective mechanism against UVR-induced DNA damage, and generally correlates inversely with the risk of skin cancer, including melanoma. Melanoma risk is also determined by susceptibility genes, one of which is the melanocortin 1 receptor (MC1R) gene. Certain MC1R alleles are strongly associated with melanoma. We hereby present experimental evidence for the role of two melanoma risk factors, constitutive pigmentation, as assessed by total melanin, eumelanin and pheomelanin contents, and MC1R genotype and function, in determining the induction and repair of DNA photoproducts in cultured human melanocytes after irradiation with increasing doses of UVR. We found that total melanin and eumelanin contents (MC and EC) correlated inversely with the extent of UVR-induced growth arrest, apoptosis and induction of cyclobutane pyrimidine dimers (CPD), but not with hydrogen peroxide release in melanocytes expressing functional MC1R. In comparison, melanocytes with loss-of-function MC1R, regardless of their MC or EC, sustained more UVR-induced apoptosis and CPD, and exhibited reduced CPD repair. Therefore, MC, mainly EC, and MC1R function are independent determinants of UVR-induced DNA damage in melanocytes.     

Biosecurity risk factors presented by international vessels: a statistical analysis

Invasive non-indigenous species are among the greatest threats to global biodiversity. Shipping is the principal vector for international dispersal of nonindigenous species, and shipping rates are increasing globally. The Australian government performs a range of regulatory actions to mitigate biosecurity risks associated with marine vessels, and in so doing has amassed a large volume of operational inspection data. This data can be used to quantitatively examine risk factors of vessels failing biosecurity procedures after arriving from international ports, the nature of biosecurity failures, and the types and seizure rates of biosecurity risk material (BRM). Classification trees with gradient boosting were used to assess characteristics that predict high risk vessels (n = 93,006) for carrying BRM, across 7 years of inspection data. Undeclared vessels and suspected irregular entry vessels posed the highest risk, but both were rare. Vessels that visit infrequently (<20 visits in 7 years) were common and had almost three times greater odds of failing inspection than vessels visiting frequently. On statistical analysis, yachts appeared to pose less risk than commercial vessels. In operational terms, a tentative profiled 20% fraction would contain 57% of genuine failures, and the concomitant non-screened group would contain 82% of passes. The most common reason for inspection failures was ballast water non-compliance (2.53%) and plant or insect detections (1.77%); biofouling was less common (0.13%) but testing for biofouling is not exhaustive. Invertebrate species comprised almost 90% of invasive organisms detected and seized from vessels failing biosecurity inspections. This study targets an entire transportation vector, which includes many pathways. Understanding the characteristics of transport vectors is pivotal to characterising the risk of biological invasions and applying adequate controls and prevention strategies. Our results show that biosecurity risk is not uniform on maritime pathways, so there is considerable scope for biosecurity regulators to impose risk-based intervention.     

Non-imidazole heterocyclic histamine H3 receptor antagonists

Continued exploration of the SAR around the lead imidazopyridine histamine H(3) antagonist 1 has led to the discovery of several related series of heterocyclic histamine H(3) antagonists. The synthesis and SAR of indolizine, indole and pyrazolopyridine based compounds are now described.     

Improved single- and multi-contact life-time testing of dental restorative materials using key characteristics of the human masticatory system and a force/position-controlled robotic dental wear simulator

This paper presents a new in vitro wear simulator based on spatial parallel kinematics and a biologically inspired implicit force/position hybrid controller to replicate chewing movements and dental wear formations on dental components, such as crowns, bridges or a full set of teeth. The human mandible, guided by passive structures such as posterior teeth and the two temporomandibular joints, moves with up to 6 degrees of freedom (DOF) in Cartesian space. The currently available wear simulators lack the ability to perform these chewing movements. In many cases, their lack of sufficient DOF enables them only to replicate the sliding motion of a single occlusal contact point by neglecting rotational movements and the motion along one Cartesian axis. The motion and forces of more than one occlusal contact points cannot accurately be replicated by these instruments. Furthermore, the majority of wear simulators are unable to control simultaneously the main wear-affecting parameters, considering abrasive mechanical wear, which are the occlusal sliding motion and bite forces in the constraint contact phase of the human chewing cycle. It has been shown that such discrepancies between the true in vivo and the simulated in vitro condition influence the outcome and the quality of wear studies. This can be improved by implementing biological features of the human masticatory system such as tooth compliance realized through the passive action of the periodontal ligament and active bite force control realized though the central nervous system using feedback from periodontal preceptors. The simulator described in this paper can be used for single- and multi-occlusal contact testing due to its kinematics and ability to exactly replicate human translational and rotational mandibular movements with up to 6 DOF without neglecting movements along or around the three Cartesian axes. Recorded human mandibular motion and occlusal force data are the reference inputs of the simulator. Experimental studies of wear using this simulator demonstrate that integrating the biological feature of combined force/position hybrid control in dental material testing improves the linearity and reduces the variability of results. In addition, it has been shown that present biaxially operated dental wear simulators are likely to provide misleading results in comparative in vitro/in vivo one-contact studies due to neglecting the occlusal sliding motion in one plane which could introduce an error of up to 49% since occlusal sliding motion D and volumetric wear loss V(loss) are proportional.     

Availability of and Ease of Access to Calorie Information on Restaurant Websites

Objective

Offering calories on restaurant websites might be particularly important for consumer meal planning, but the availability of and ease of accessing this information are unknown.

Methods

We assessed websites for the top 100 U.S. chain restaurants to determine the availability of and ease of access to calorie information as well as website design characteristics. We also examined potential predictors of calorie availability and ease of access.

Results

Eighty-two percent of restaurants provided calorie information on their websites; 25% presented calories on a mobile-formatted website. On average, calories could be accessed in 2.35±0.99 clicks. About half of sites (51.2%) linked to calorie information via the homepage. Fewer than half had a separate section identifying healthful options (46.3%), or utilized interactive meal planning tools (35.4%). Quick service/fast casual, larger restaurants, and those with less expensive entrées and lower revenue were more likely to make calorie information available. There were no predictors of ease of access.

Conclusion

Calorie information is both available and largely accessible on the websites of America’s leading restaurants. It is unclear whether consumer behavior is affected by the variability in the presentation of calorie information.     

Betaine and Trimethylamine-N-Oxide as Predictors of Cardiovascular Outcomes Show Different Patterns in Diabetes Mellitus: An Observational Study

Background

Betaine is a major osmolyte, also important in methyl group metabolism. Concentrations of betaine, its metabolite dimethylglycine and analog trimethylamine-N-oxide (TMAO) in blood are cardiovascular risk markers. Diabetes disturbs betaine: does diabetes alter associations between betaine-related measures and cardiovascular risk?

Methods

Plasma samples were collected from 475 subjects four months after discharge following an acute coronary admission. Death (n = 81), secondary acute MI (n = 87), admission for heart failure (n = 85), unstable angina (n = 72) and all cardiovascular events (n = 283) were recorded (median follow-up: 1804 days).

Results

High and low metabolite concentrations were defined as top or bottom quintile of the total cohort. In subjects with diabetes (n = 79), high plasma betaine was associated with increased frequencies of events; significantly for heart failure, hazard ratio 3.1 (1.2–8.2) and all cardiovascular events, HR 2.8 (1.4–5.5). In subjects without diabetes (n = 396), low plasma betaine was associated with events; significantly for secondary myocardial infarction, HR 2.1 (1.2–3.6), unstable angina, HR 2.3 (1.3–4.0), and all cardiovascular events, HR 1.4 (1.0–1.9). In diabetes, high TMAO was a marker of all outcomes, HR 2.7 (1.1–7.1) for death, 4.0 (1.6–9.8) for myocardial infarction, 4.6 (2.0–10.7) for heart failure, 9.1 (2.8–29.7) for unstable angina and 2.0 (1.1–3.6) for all cardiovascular events. In subjects without diabetes TMAO was only significant for death, HR 2.7 (1.6–4.8) and heart failure, HR 1.9 (1.1–3.4). Adding the estimated glomerular filtration rate to Cox regression models tended to increase the apparent risks associated with low betaine.

Conclusions

Elevated plasma betaine concentration is a marker of cardiovascular risk in diabetes; conversely low plasma betaine concentrations indicate increased risk in the absence of diabetes. We speculate that the difference reflects control of osmolyte retention in tissues. Elevated plasma TMAO is a strong risk marker in diabetes.     

Novel tetrahydroisoquinolines are histamine H3 antagonists and serotonin reuptake inhibitors

A series of novel 4-aryl-1,2,3,4-tetrahydroisoquinoline-based histamine H(3) ligands that also have serotonin reuptake transporter inhibitor activity is described. The synthesis, in vitro biological data, and select pharmacokinetic data for these novel compounds are discussed.     

Identification of a putative acetyltransferase gene, MMP0350, which affects proper assembly of both flagella and pili in the archaeon Methanococcus maripaludis

Glycosylation is a posttranslational modification utilized in all three domains of life. Compared to eukaryotic and bacterial systems, knowledge of the archaeal processes involved in glycosylation is limited. Recently, Methanococcus voltae flagellin proteins were found to have an N-linked trisaccharide necessary for proper flagellum assembly. Current analysis by mass spectrometry of Methanococcus maripaludis flagellin proteins also indicated the attachment of an N-glycan containing acetylated sugars. To identify genes involved in sugar biosynthesis in M. maripaludis, a putative acetyltransferase was targeted for in-frame deletion. Deletion of this gene (MMP0350) resulted in a flagellin molecular mass shift to a size comparable to that expected for underglycosylated or completely nonglycoslyated flagellins, as determined by immunoblotting. Assembled flagellar filaments were not observed by electron microscopy. Interestingly, the deletion also resulted in defective pilus anchoring. Mutant cells with a deletion of MMP0350 had very few, if any, pili attached to the cell surface compared to a nonflagellated but piliated strain. However, pili were obtained from culture supernatants of this strain, indicating that the defect was not in pilus assembly but in stable attachment to the cell surface. Complementation of MMP0350 on a plasmid restored pilus attachment, but it was unable to restore flagellation, likely because the mutant ceased to make detectable flagellin. These findings represent the first report of a biosynthetic gene involved in flagellin glycosylation in archaea. Also, it is the first gene to be associated with pili, linking flagellum and pilus structure and assembly through posttranslational modifications.