Long-Term Quantitative Evaluation of Muscle and Bone Wasting Induced by Botulinum Toxin in Mice Using Microcomputed Tomography.

Muscle and bone masses are highly correlated and muscles impose large loads on bone. Muscle wasting that accompanies bone loss has been poorly investigated. 21 female mice were spread into seven groups. At day 0, 18 mice received Botulinum toxin (BTX) injection in the quadriceps muscle to induce paralysis of the right hind limb; the left contralateral side was used as control. Mice were sacrificed at 7, 14, 21, 28, 56 and 90 days post-injection. A remaining group was sacrificed at day 0. Trabecular bone volume was determined by microcomputed tomography (microCT) at the distal femur and tibia proximal metaphyses on both sides. Limbs were immersed in an HgCl2 solution allowing muscle visualization by microCT. On 2D sections, the cross-sectional areas and form-factors were measured for the quadriceps at mid-thigh and gastrocnemius at mid-leg and these muscles were dissected and weighed. Bone volume decreased in the paralysed side. Bone loss was maximal at 56 days followed by recuperation at 90 days. The cross-sectional areas of gastrocnemius and quadriceps were significantly lower in the paralysed limb from 7 days; the decrease was maximum at 21 days for the gastrocnemius and 28 days for the quadriceps. No difference in form-factors was found between the two limbs. Similar results were obtained with the anatomical method and significant correlations were obtained between the two methods. Quantitative analysis of muscle loss and recovery was possible by microCT after using a metallic contrast agent. Loss of bone secondary to muscle wastage induced by BTX and recovery showed a parallel evolution for bone and muscles.

Development and evaluation of a simple treatment eligibility score (HEPSANET) to decentralise hepatitis B care in Africa: a cross-sectional study.

BACKGROUND: Hepatitis B virus (HBV) elimination requires expanding and decentralising HBV care services. However, peripheral health facilities lack access to diagnostic tools to assess eligibility for antiviral therapy. Through the Hepatitis B in Africa Collaborative Network (HEPSANET), we aimed to develop and evaluate a score using tests generally available at lower-level facilities, to simplify the evaluation of antiviral therapy eligibility in people living with HBV. METHODS: We surveyed the availability of clinical and laboratory parameters across different health-care levels in sub-Saharan Africa. We used data from the HEPSANET dataset, the largest cross-sectional dataset of treatment-naive people living with HBV in sub-Saharan Africa, to derive and validate the score. Participants from this dataset were included in the analysis if they were aged 18 years or older and had liver fibrosis stages determined by a liver stiffness measurement or liver histopathology. Participants with co-infections or metabolic disorders were excluded. We allocated participants to the derivation and validation sets by geographical site. In the derivation set, we used stepwise logistic regression to identify the best performing parameters for identifying participants that met the 2017 European Association for the Study of the Liver (EASL) criteria. Regression coefficients were converted into integer points to construct simplified algorithms for different health-care levels. In the validation set, we estimated the area under the receiver operating characteristic, sensitivity, and specificity of the simplified algorithm for identifying antiviral therapy eligibility defined by the 2017 EASL criteria. FINDINGS: At 11 sites from eight countries that returned surveys, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and platelet count were generally available at district hospital levels, and hepatitis B e antigen and point-of-care HBV DNA tests were available only at regional and provincial hospital levels or above. Among 2895 participants included from the HEPSANET database (1740 [60·1%] male, 1155 [39·9%] female), 409 (14·1%) met EASL antiviral therapy eligibility criteria. In the derivation set, the optimal district-level hospital score was: ALT (IU/L), less than 40 (0 points), 40-79 (+1), 80 or greater (+2); AST (IU/L), less than 40 (0), 40-79 (+1), 80 or greater (+2); and platelet counts (109/L), less than 100 (+2), 100-149 (+1), 150 or greater (0). When combined with family history and clinical data for decompensated cirrhosis that do not require any biological tests, a cut-off of 2 points or more had a sensitivity and specificity of 82% (95% CI 76-86) and 95% (93-96) to identify treatment-eligible individuals in the derivation set, and 78% (71-85) and 87% (86-89) in the validation set, respectively. INTERPRETATION: Using a score incorporating platelet counts, AST, and ALT, the majority of people living with HBV requiring antiviral therapy can be identified. Our findings suggest that clinical staging can be decentralised down to district hospital level in sub-Saharan Africa. FUNDING: European Association for the Study of the Liver Foundation, John C Martin Foundation. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Recapitulation of Human Embryonic Heartbeat to Promote Differentiation of Hepatic Endoderm to Hepatoblasts.

Hepatic development requires multiple sequential physicochemical environmental changes in an embryo, and human pluripotent stem cells (hPSCs) allow for the elucidation of this embryonic developmental process. However, the current in vitro methods for hPSC-hepatic differentiation, which employ various biochemical substances, produce hPSC-derived hepatocytes with less functionality than primary hepatocytes, due to a lack of physical stimuli, such as heart beating. Here, we developed a microfluidic platform that recapitulates the beating of a human embryonic heart to improve the functionality of hepatoblasts derived from hepatic endoderm (HE) in vitro. This microfluidic platform facilitates the application of multiple mechanical stretching forces, to mimic heart beating, to cultured hepatic endoderm cells to identify the optimal stimuli. Results show that stimulated HE-derived hepatoblasts increased cytochrome P450 3A (CYP3A) metabolic activity, as well as the expression of hepatoblast functional markers (albumin, cytokeratin 19 and CYP3A7), compared to unstimulated hepatoblasts. This approach of hepatic differentiation from hPSCs with the application of mechanical stimuli will facilitate improved methods for studying human embryonic liver development, as well as accurate pharmacological testing with functional liver cells.

Biophysical Control of Bile Duct Epithelial Morphogenesis in Natural and Synthetic Scaffolds.

The integration of bile duct epithelial cells (cholangiocytes) in artificial liver culture systems is important in order to generate more physiologically relevant liver models. Understanding the role of the cellular microenvironment on differentiation, physiology, and organogenesis of cholangiocytes into functional biliary tubes is essential for the development of new liver therapies, notably in the field of cholangiophaties. In this study, we investigated the role of natural or synthetic scaffolds on cholangiocytes cyst growth, lumen formation and polarization. We demonstrated that cholangiocyte cyst formation efficiency can be similar between natural and synthetic matrices provided that the mechanical properties of the hydrogels are matched. When using synthetic matrices, we also tried to understand the impact of elasticity, matrix metalloprotease-mediated degradation and integrin ligand density on cyst morphogenesis. We demonstrated that hydrogel stiffness regulates cyst formation. We found that controlling integrin ligand density was key in the establishment of large polarized cysts of cholangiocytes. The mechanism of lumen formation was found to rely on cell self-organization and proliferation. The formed cholangiocyte organoids showed a good MDR1 (multi drug resistance protein) transport activity. Our study highlights the advantages of fully synthetic scaffold as a tool to develop bile duct models.