RESUMEN
The development of inflammatory bowel diseases (IBD) involves the breakdown of two barriers: the epithelial barrier and the gut-vascular barrier (GVB). The destabilization of each barrier can promote initiation and progression of the disease. Interestingly, first evidence is available that both barriers are communicating through secreted factors that may accordingly serve as targets for therapeutic modulation of barrier functions. Interferon (IFN)-γ is among the major pathogenesis factors in IBD and can severely impair both barriers. In order to identify factors transmitting signals from the GVB to the epithelial cell barrier, we analyzed the secretome of IFN-γ-treated human intestinal endothelial cells (HIEC). To this goal, HIEC were isolated in high purity from normal colon tissues. HIEC were either untreated or stimulated with IFN-γ (10 U/mL). After 48 h, conditioned media (CM) were harvested and subjected to comparative hyper reaction monitoring mass spectrometry (HRM™ MS). In total, 1,084 human proteins were detected in the HIEC-CM. Among these, 43 proteins were present in significantly different concentrations between the CM of IFN-γ- and control-stimulated HIEC. Several of these proteins were also differentially expressed in various murine colitis models as compared to healthy animals supporting the relevance of these proteins secreted by inflammatory activated HIEC in the inter-barrier communication in IBD. The angiocrine pathogenic impact of these differentially secreted HIEC proteins on the epithelial cell barrier and their perspectives as targets to treat IBD by modulation of trans-barrier communication is discussed in detail.
RESUMEN
Three-Dimensional (3D) liver microtissues, specifically prepared from primary human hepatocytes (PHH) in coculture with nonparenchymal cells (NPCs), have been shown to be a valuable tool for in vitro toxicology. However, a lack of thorough characterization on a functional, transcriptomic, and proteomic level of such models during long-term cultivation is evident. By integrating multiple omics technologies, we provide in this study an in-depth long-term characterization of 3D microtissues composed of PHH from three different donors cocultured with primary NPCs. The 3D human liver microtissues (hLiMTs) exhibited stable adenosine triphosphate (ATP) content and albumin secretion over 5 weeks. Histological analysis indicated a healthy liver tissue with polarized expression of bile salt export pump (BSEP) and multidrug resistance protein 2 (MRP2) in a structure reminiscent of bile canaliculi. The 3D microtissues exhibited stable basal and inducible cytochrome P450 activities up to 5 weeks in culture. Analysis of 40,716 transcripts using RNA arrays revealed distinct similarities to native human liver gene expression. Long-term culture showed a stable phenotype up to 5 weeks, with differences in liver gene expression primarily attributed to individual donors. Proteomic profiling of 2200 unique proteins by label-free LC-MS/MS revealed a relatively stable protein expression where only 7.3% were up- or downregulated more than twofold from day 7 to 35 in culture. Taken together, these results suggest that hLiMTs represent a responsive and physiologically relevant in vitro liver model that maintains stable function over 5 weeks and is therefore well suited for repeated-dose toxicity testing.
RESUMEN
INTRODUCTION: The beneficial effect of obesity on bone mineral density (BMD) has not been definitely established. OBJECTIVES: The aim of the study was to evaluate changes in BMD in obese perimenopausal women during a 5-year follow-up. PATIENTS AND METHODS: The study involved 54 obese women. The group was divided into 2 subgroups according to the menopausal status: postmenopausal women--M (n = 35) and premenopausal women--P (n = 19). Laboratory tests (parathyroid hormone, 25-hydroxyvitamin D3, C-terminal telopeptide of type I collagen, osteocalcin, and osteoprotegerin), anthropometric measurements, and densitometry were performed twice during the 5-year follow-up. The control group consisted of 19 healthy women of the same age and with normal body weight. RESULTS: Obese postmenopausal women were characterized by lower BMD in the proximal femur and lumbar spine, higher fracture risk, and higher serum osteocalcin levels at baseline. During the 5-year follow-up, there was a 1.52% and 6.86% decrease in proximal femur BMD and 2.34% and 5.17% in lumbar spine BMD (in premenopausal and postmenopausal women, respectively). In postmenopausal controls, BMD reduction was 2.36% and 4.3%, respectively. In the combined analysis including all postmenopausal women, there was an inverse correlation between the initial body mass index and the changes in proximal femur BMD (r = -0.25; P <0.05) and lumbar spine BMD (r = -0.28; P = 0.08) that occurred during the 5-year follow-up. CONCLUSIONS: Obesity appears not to protect against bone mineral loss in postmenopausal women.