Dchs1-Fat4 regulation of osteogenic differentiation in mouse.

In human, mutations of the protocadherins FAT4 and DCHS1 result in Van Maldergem syndrome, which is characterised, in part, by craniofacial abnormalities. Here, we analyse the role of Dchs1-Fat4 signalling during osteoblast differentiation in mouse. We show that Fat4 and Dchs1 mutants mimic the craniofacial phenotype of the human syndrome and that Dchs1-Fat4 signalling is essential for osteoblast differentiation. In Dchs1/Fat4 mutants, proliferation of osteoprogenitors is increased and osteoblast differentiation is delayed. We show that loss of Dchs1-Fat4 signalling is linked to increased Yap-Tead activity and that Yap is expressed and required for proliferation in osteoprogenitors. In contrast, Taz is expressed in more-committed Runx2-expressing osteoblasts, Taz does not regulate osteoblast proliferation and Taz-Tead activity is unaffected in Dchs1/Fat4 mutants. Finally, we show that Yap and Taz differentially regulate the transcriptional activity of Runx2, and that the activity of Yap-Runx2 and Taz-Runx2 complexes is altered in Dchs1/Fat4 mutant osteoblasts. In conclusion, these data identify Dchs1-Fat4 as a signalling pathway in osteoblast differentiation, reveal its crucial role within the early Runx2 progenitors, and identify distinct requirements for Yap and Taz during osteoblast differentiation.

Periapical status in patients affected by osteoporosis: A retrospective clinical study.

OBJECTIVES: To assess the periapical status in patients with osteoporosis (OP) treated with denosumab (D), bisphosphonates (BPs), or not on medication, and to understand if these conditions influence the prevalence and the progression of apical periodontitis (AP). MATERIAL AND METHODS: Seventy-six patients with OP alone or treated with D, or BPs, formed the study group (O), and those from 76 patients matched for age and sex, without diseases, and not taking medications, constituted the control (C) in this retrospective case-control study. The data from the complete clinical and radiographic examination, medical history, decayed, missing, and filled teeth (DMFT), and periapical index score (PAI) were recorded for each patient. Wilcoxon rank test, χ2 , and Student's t test were used as appropriate. RESULTS: The prevalence of AP was similar in O and C. Furthermore, AP was significantly more frequent in root canal-treated teeth in O patients (p = .03). CONCLUSIONS: OP does not appear to be associated with the development of AP. Moreover, the increased prevalence of AP in root canal-treated teeth in O patients highlights a possible relationship between the healing dynamics of the disease post-therapy and the patients' medication. A larger sample is needed to confirm these findings.

Evidence of Amniotic Epithelial Cell Differentiation toward Hepatic Sinusoidal Endothelial Cells.

Amniotic epithelial cells (AECs) represent a useful and noncontroversial source for liver-based regenerative medicine, as they can differentiate into hepatocytes upon transplantation into the liver. However, the possibility that AECs can differentiate into other liver cell types, such as hepatic sinusoidal endothelial cells (HSECs), has never been assessed. In order to test this hypothesis, rat- and human-derived AECs (rAECs and hAECs, respectively) were subjected to endothelial cell tube formation assay in vitro. Moreover, to evaluate differentiation in vivo, the retrorsine (RS) model of liver repopulation was used. Pyrrolizidine alkaloids (including RS) are known to target both hepatocytes and endothelial cells, inducing cell enlargement and inhibition of cell cycle progression. rAECs and hAECs were able to form capillary-like structures when cultured under proangiogenic conditions. For in vivo experiments, rAECs were obtained from dipeptidyl peptidase type IV (DPP-IV, CD26) donors and were transplanted into the liver of recipient CD26 negative animals pretreated with RS. rAEC-derived cells were engrafted in between hepatocytes and resembled HSECs as assessed by morphological analysis and the pattern of expression of CD26. Donor-derived CD26+ cells coexpressed HSEC markers RECA-1 and SE-1, while they lacked expression of typical hepatocyte markers (i.e., cytochrome P450, hepatocyte nuclear factor 4α). As such, these results provide the first evidence that AECs can respond to proangiogenic signals in vitro and differentiate into HSECs in vivo. Furthermore, they support the conclusion that AECs possesses great plasticity and represents a promising tool in the field of regenerative medicine both in the liver and in other organs.

Caloric restriction promotes rapid expansion and long-lasting increase of Lactobacillus in the rat fecal microbiota.

Previous studies indicated that caloric restricted diet enables to lower significantly the risk of cardiovascular and metabolic diseases. In experimental animal models, life-long lasting caloric restriction (CR) was demonstrated to induce changes of the intestinal microbiota composition, regardless of fat content and/or exercise. To explore the potential impact of short and long-term CR treatment on the gut microbiota, we conducted an analysis of fecal microbiota composition in young and adult Fisher 344 rats treated with a low fat feed under ad libitum (AL) or CR conditions (70%). We report here significant changes of the rat fecal microbiota that arise rapidly in young growing animals after short-term administration of a CR diet. In particular, Lactobacillus increased significantly after 8 weeks of CR treatment and its relative abundance was significantly higher in CR vs AL fed animals after 36 weeks of dietary intervention. Taken together, our data suggest that Lactobacillus intestinal colonization is hampered in AL fed young rats compared to CR fed ones, while health-promoting CR diet intervention enables the expansion of this genus rapidly and persistently up to adulthood.

Caloric restriction delays early phases of carcinogenesis via effects on the tissue microenvironment.

Caloric restriction (CR) is an effective and consistent means to delay aging and the incidence of chronic diseases related to old age, including cancer. However, the precise mechanisms responsible for the beneficial effect of CR on carcinogenic process are yet to be identified.In the present studies the hypothesis was tested that the CR might delay carcinogenesis via modulatory effects exerted on the age-associated, neoplastic-prone tissue microenvironment. Using a well characterized, orthotopic cell transplantation (Tx) system in the rat, preneoplastic hepatocytes isolated from liver nodules were injected into either old syngeneic rats fed ad libitum (AL) or animals of the same age given a CR diet (70% of AL feeding). Analysis of donor-derived cell clusters performed at 10 weeks post-Tx revealed a significant shift towards smaller class sizes in the group receiving CR diet. Clusters comprising more than 50 cells, including large hepatic nodules, were thrice more frequent in AL vs. CR animals. Incidence of spontaneous endogenous nodules was also decreased by CR. Markers of cell senescence were equally expressed in the liver of AL and CR groups. However, higher levels of SIRT1 and FOXO1 proteins were detected in CR-exposed livers, while expression of HDAC1 and C/EBPß were decreased. These results are interpreted to indicate that CR delays the emergence of age-associated neoplastic disease through effects exerted, at least in part, on the tissue microenvironment. Nutrient-sensing pathways might mediate such modulatory effect.

A GC-MS untargeted metabolomics analysis in the plasma and liver of rats lacking dipeptidyl-peptidase type IV enzyme activity.

This study was achieved with the aim to find metabolic changes between Fischer rats with different dipeptidyl peptidase-type 4 (DPPIV) expression. The DPPIV is an enzyme expressed in several tissues and is critically involved in the regulation of meal-related insulin secretion in healthy individuals. The metabolic consequences of chronic DPPIV inhibition were analyzed in a surrogate animal model of genetic enzyme deficiency. Hyphenated gas chromatography-mass spectrometry (GC-MS) and multivariate data analysis techniques were used to study the metabolic aqueous fraction profile of 18 plasma and liver samples in two syngeneic rat strains differing in DPPIV activity (DPPIV+ vs. DPPIV-). The hyperglycemic response following oral glucose administration was attenuated in DPPIV- rats, as expected. Statistical significant differences between the two strains were observed among the low molecular weight polar metabolites analyzed from plasma and liver.These included a decrease in malic acid and glutamine and an increase in pyroglutamic acid, serine, and alanine in plasma of DPPIV- rats. In addition, palmitic acid, L-proline, and ribitol were decreased in the liver of DPPIV- strain. Such alterations were compatible with a normal phenotype. These results suggest that long-term exposure to DPPIV inhibitors looks compatible with an overall balanced metabolism.

Differentiation of amniotic epithelial cells into various liver cell types and potential therapeutic applications.

The aim of Regenerative Medicine is to replace or regenerate human cells, tissues or organs in order to restore normal function. Among all organs, the liver is endowed with remarkable regenerative capacity. Nonetheless, there are conditions in which this ability is impaired, and the use of isolated cells, including stem cells, is being considered as a possible therapeutic tool for the management of chronic hepatic disease. Placenta holds great promise for the field of regenerative medicine. It has long been used for the treatment of skin lesions and in ophthalmology, due to its ability to modulate inflammation and promote healing. More recently, cells isolated from the amniotic membrane are being considered as a possible resource for tissue regeneration, including in the context liver disease. Two cell types can be easily isolated from human amnion: epithelial cells (hAEC) and mesenchymal stromal cells (hAMSC). However only the first cell population has been demonstrated to be a possible source of proficient hepatic cells. This review will summarize current knowledge on the differentiation of hAEC into liver cells and their potential therapeutic application.

Hyperplasia vs hypertrophy in tissue regeneration after extensive liver resection.

AIM: To address to what extent hypertrophy and hyperplasia contribute to liver mass restoration after major tissue loss. METHODS: The ability of the liver to regenerate is remarkable on both clinical and biological grounds. Basic mechanisms underlying this process have been intensively investigated. However, it is still debated to what extent hypertrophy and hyperplasia contribute to liver mass restoration after major tissue loss. We addressed this issue using a genetically tagged system. We were able to follow the fate of single transplanted hepatocytes during the regenerative response elicited by 2/3 partial surgical hepatectomy (PH) in rats. Clusters of transplanted cells were 3D reconstructed and their size distribution was evaluated over time after PH. RESULTS: Liver size and liver DNA content were largely recovered 10 d post-PH, as expected (e.g., total DNA/liver/100 g b.w. was 6.37 ± 0.21 before PH and returned to 6.10 ± 0.36 10 d after PH). Data indicated that about 2/3 of the original residual hepatocytes entered S-phase in response to PH. Analysis of cluster size distribution at 24, 48, 96 h and 10 d after PH revealed that about half of the remnant hepatocytes completed at least 2 cell cycles. Average size of hepatocytes increased at 24 h (248.50 µm2 ± 7.82 µm2, P = 0.0015), but returned to control values throughout the regenerative process (up to 10 d post-PH, 197.9 µm2 ± 6.44 µm2, P = 0.11). A sizeable fraction of the remnant hepatocyte population does not participate actively in tissue mass restoration. CONCLUSION: Hyperplasia stands as the major mechanism contributing to liver mass restoration after PH, with hypertrophy playing a transient role in the process.

Regenerative Medicine: Shedding Light on the Link between Aging and Cancer.

The evidence linking aging and cancer is overwhelming. Findings emerging from the field of regenerative medicine reinforce the notion that aging and cancer are profoundly interrelated in their pathogenetic pathways. We discuss evidence to indicate that age-associated alterations in the tissue microenvironment contribute to the emergence of a neoplastic-prone tissue landscape, which is able to support the selective growth of preneoplastic cell populations. Interestingly, tissue contexts that are able to select for the growth of preneoplastic cells, including the aged liver microenvironment, are also supportive for the clonal expansion of normal, homotypic, transplanted cells. This suggests that the growth of normal and preneoplastic cells is possibly driven by similar mechanisms, implying that strategies based on principles of regenerative medicine might be applicable to modulate neoplastic disease.

Aging promotes neoplastic disease through effects on the tissue microenvironment.

A better understanding of the complex relationship between aging and cancer will provide important tools for the prevention and treatment of neoplasia. In these studies, the hypothesis was tested that aging may fuel carcinogenesis via alterations imposed in the tissue microenvironment. Preneoplastic hepatocytes isolated from liver nodules were orthotopically injected into either young or old syngeneic rats and their fate was followed over time using the dipeptidyl-peptidase type IV (DPPIV) system to track donor-derived-cells. At 3 months post-Tx, the mean size of donor-derived clusters was 11±3 cells in young vs. 42±8 in old recipients. At 8 months post-Tx, no visible lesion were detected in any of 21 young recipients, while 17/18 animals transplanted at old age displayed hepatic nodules, including 7 large tumors. All tumors expressed the DPPIV marker enzyme, indicating that they originated from transplanted cells. Expression of senescence-associated ß-galactosidase was common in liver of 18-month old animals, while it was a rare finding in young controls. Finally, both mRNA and IL6 protein were found to be increased in the liver of aged rats compared to young controls. These results are interpreted to indicate that the microenvironment of the aged liver promotes the growth of pre-neoplastic hepatocytes.

A GC-MS untargeted metabolomics analysis in the plasma and liver of rats lacking dipeptidyl-peptidase type IV enzyme activity

This study was achieved with the aim to find metabolic changes between Fischer rats with different dipeptidyl peptidase-type 4 (DPPIV) expression. The DPPIV is an enzyme expressed in several tissues and is critically involved in the regulation of meal-related insulin secretion in healthy individuals. The metabolic consequences of chronic DPPIV inhibition were analyzed in a surrogate animal model of genetic enzyme deficiency. Hyphenated gas chromatography–mass spectrometry (GC-MS) and multivariate data analysis techniques were used to study the metabolic aqueous fraction profile of 18 plasma and liver samples in two syngeneic rat strains differing in DPPIV activity (DPPIV+ vs. DPPIV-). The hyperglycemic response following oral glucose administration was attenuated in DPPIV- rats, as expected. Statistical significant differences between the two strains were observed among the low molecular weight polar metabolites analyzed from plasma and liver. These included a decrease in malic acid and glutamine and an increase in pyroglutamic acid, serine, and alanine in plasma of DPPIV- rats. In addition, palmitic acid, l-proline, and ribitol were decreased in the liver of DPPIV- strain. Such alterations were compatible with a normal phenotype. These results suggest that long-term exposure to DPPIV inhibitors looks compatible with an overall balanced metabolism