Optimization of 3-D hepatocyte culture by controlling the physical and chemical properties of the extra-cellular matrices

Hepatocytes are anchorage-dependent cells sensitive to microenvironment; the control of the physicochemical properties of the extra-cellular matrices may be useful to the maintenance of hepatocyte functions in vitro for various applications. In a microcapsule-based 3-D hepatocyte culture microenvironment, we could control the physical properties of the collagen nano-fibres by fine-tuning the complex-coacervation reaction between methylated collagen and terpolymer of hydroxylethyl methacrylate-methyl methacrylate-methylacrylic acid. The physical properties of the nano-fibres were quantitatively characterized using back-scattering confocal microscopy to help optimize the physical support for hepatocyte functions. We further enhanced the chemical properties of the collagen nano-fibres by incorporating galactose onto collagen, which can specifically interact with the asialoglycoprotein receptor on hepatocytes. By correlating a range of collagen nano-fibres of different physicochemical properties with hepatocyte functions, we have identified a specific combination of methylated and galactosylated collagen nano-fibres optimal for maintaining hepatocyte functions in vitro. A model of how the physical and chemical supports interplay to maintain hepatocyte functions is discussed.     

Adhesion contact dynamics of HepG2 cells on galactose-immobilized substrates

The specific recognition between asialoglycoprotein receptor and galactose ligand at cell-substrate interfaces has been shown to mediate hepatocyte adhesion and maintain liver specific functions of hepatocytes. Conventionally, the success of hepatocyte attachment on engineered tissue scaffold is inferred from the degree of two-dimensional cell spreading that is measured by transmitted light microscopy. However, the actual contact mechanics and adhesion strength of hepatocytes during two-dimensional cell spreading has not been elucidated due to lack of biophysical probe. In this study, a novel biophysical technique known as confocal reflectance interference contrast microscopy (C-RICM) in conjunction with phase contrast microscopy is utilized to probe the adhesion dynamics, contact mechanics and two-dimensional spreading kinetics of HepG2 cells on galactose immobilized and collagen gel coated substrates. C-RICM demonstrates that HepG2 cells form strong adhesion contacts with both galactose-immobilized surfaces and collagen gel coated substrates. Moreover, HepG2 cells maintain their compact shapes in the presence of asialoglycoprotein receptor-mediated recognition while they become exceedingly spread under integrin-mediated adhesion on collagen gel coated substrate. The initial rate of adhesion contact formation and the steady-state adhesion energy of HepG2 cell population are highest on substrate conjugated with galactose ligand via a longer spacer. The adhesion dynamics and final adhesion energy of HepG2 cells depends both on the type of ligand-receptor interaction and the length of spacer between the ligand and substrate. Most importantly, new biophysical insights into the initial hepatocyte attachment that are critical for hepatocyte culture are provided through the decomposition of two-dimensional spreading and adhesion contact formation on bio-functional substrates.     

Galactosylated PVDF membrane promotes hepatocyte attachment and functional maintenance

One of the major challenges in BLAD design is to develop functional substrates suitable for hepatocyte attachment and functional maintenance. In the present study, we designed a poly(vinylidene difluoride) (PVDF) surface coated with galactose-tethered Pluronic polymer. The galactose-derived Pluronic F68 (F68-Gal) was adsorbed on PVDF membrane through hydrophobic-hydrophobic interaction between PVDF and the polypropylene oxide segment in Pluronic. The galactose density on the modified PVDF surface increased with the concentration of the F68-Gal solution, reaching 15.4 nmol galactosyl groups per cm2 when a 1 mg/ml of F68-Gal solution was used. The adsorbed F68-Gal remained relatively stable in culture medium. Rat hepatocytes attachment efficiency on F68-Gal modified PVDF membrane was similar to that on collagen-coated surface. The attached hepatocytes on PVDF/F68-Gal membrane self-assembled into multi-cellular spheroids after 1 day of culture. These attached hepatocytes in spheroids exhibited higher cell functions such as albumin synthesis and P450 1A1 detoxification function compared to unmodified PVDF membrane and collagen-coated surface. These results suggest the potential of this galactose-immobilized PVDF membrane as a suitable substrate for hepatocyte culture.     

Coping strategies,hostility, and depressive symptoms: A path model

Previous studies of coping, hostility, and depressive symptoms have highlighted the significant relations between all possible pairs of these 3 variables. To more completely explore the nature of depressive symptoms, we link them all together in this study by testing a coping→hostility→depressive symptoms path model. One hundred forty participants completed psychological questionnaires measuring coping strategies, hostility, and depressive symptoms. While controlling age and social class as covariates, SPSS stepwise regression analyses were used to examine relations among these 3 constructs. Results suggest that coping has a direct relation with depressive symptoms as well as an indirect relation mediated by hostility. Passive coping may lead to increased hostility, resulting in depressive symptoms. Active coping may have the opposite effect. These findings suggest that the inclusion of measures of both coping strategies and hostility yields a more thorough understanding of concomitants of depressive symptoms. From a clinical perspective, knowing what coping strategies a person uses and how much anger they experience and express may be useful in guiding the management of depressive symptoms.     

Effect of aging on neuroglobin expression in rodent brain

Neuroglobin (Ngb), a recently discovered O2-binding heme protein related to hemoglobin and myoglobin, protects neurons from hypoxic-ischemic injury in vitro and in vivo. In immunostained mouse brain sections, we found widespread expression of Ngb protein in neurons, but not astrocytes, of several brain regions that are prominently involved in age-related neurodegenerative disorders. Western blots from young adult (3 month), middle-aged (12 month), and aged (24 month) rats showed an age-related decline in Ngb expression in cerebral neocortex, hippocampus, caudate-putamen, and cerebellum. Loss of this neuroprotective protein may have a role in increasing susceptibility to age-related neurological disorders.     

Radioprotective Effect of Whey Hydrolysate Peptides against γ-Radiation-Induced Oxidative Stress in BALB/c Mice

Radiation therapy is widely used in the treatment of tumor diseases, but it can also cause serious damage to the body, so it is necessary to find effective nutritional supplements. The main purpose of this study is to evaluate the protective effect of whey hydrolysate peptides (WHPs) against ⁶⁰Coγ radiation damage in mice and explore the mechanism. BALB/c mice were given WHPs by oral gavage administration for 14 days. Then, some mice underwent a 30-day survival test after 8 Gy radiation, and other mice received 3.5 Gy radiation to analyze the changes in body weight, hematology and bone marrow DNA after three and 14 days. In addition, through further analysis of the level of oxidative stress and intestinal barrier function, the possible mechanism of the radioprotective effect of WHPs was explored. The study found WHPs can prolong survival time, restore body weight, and increase the number of peripheral blood white blood cells and bone marrow DNA content in irradiated mice. In addition, WHPs can significantly improve the antioxidant capacity, inhibit pro-inflammatory cytokines and protect the intestinal barrier. These results indicate that WHPs have a certain radioprotective effect in mice, and the main mechanism is related to reducing oxidative damage.     

Clinicopathological characteristics,prognosis, and chemosensitivity in patients with metastatic upper tract urothelial carcinoma

PurposeTo investigate the clinical characteristics, chemosensitivity, and outcome of metastatic upper tract urothelial carcinoma (UTUC).Patients and MethodsRecords of patients with metastatic UTUC since January 2005 were retrieved from a database that included clinical and survival data. Statistical analyses including survival and multivariate analyses of factors were respectively performed by the Kaplan-Meier method and Cox proportional hazard model.ResultsA total of 250 consecutive UTUC cases were evaluated. There were 56 patients (22.4%) with initially diagnosed stage IV disease. The most common metastatic sites were lung (39.6%), distant lymph nodes (39.2%), bone (19.6%), liver (18.0%), and adrenal gland (7.2%), respectively, and the local recurrence rate was 10.4%. Two hundred thirteen patients received first-line chemotherapy. The overall response rate was only 28.7% and the median progression-free survival time was only 5.0 months. The overall survival time of the cohort was 18.0 months. Multivariate analyses showed that initially diagnosed stage IV disease, number of metastatic organs ≥3, no response to chemotherapy and cycles of chemotherapy ≤2 were adverse prognosticators for overall survival.ConclusionUTUC presented to be more prone to metastasize than locally recur and thought to have low chemosensitivity. Stage IV disease at initial diagnosis, number of metastatic organs, response and cycles of chemotherapy were independent prognosticators for metastatic UTUC.     

Changing Levels of Social Engagement with Gay Men Is Associated with HIV Related Outcomes and Behaviors: Trends in Australian Behavioral Surveillance 1998–2020

Archives of Sexual Behavior - Changes to how gay, bisexual, and other men who have sex with men (GBM) connect with each other and with their communities have implications for HIV prevention. Social...