Mmp2 Deficiency Leads to Defective Parturition and High Dystocia Rates in Mice.

Parturition is the final and essential step for mammalian reproduction. While the uterus is quiescent during pregnancy, fundamental changes arise in the myometrial contractility, inducing fetal expulsion. Extracellular matrix (ECM) remodeling is fundamental for these events. The gelatinases subgroup of matrix metalloproteinases (MMPs), MMP2 and MMP9, participate in uterine ECM remodeling throughout pregnancy and parturition. However, their loss-of-function effect is unknown. Here, we determined the result of eliminating Mmp2 and/or Mmp9 on parturition in vivo, using single- and double-knockout (dKO) mice. The dystocia rates were measured in each genotype, and uterine tissue was collected from nulliparous synchronized females at the ages of 2, 4, 9 and 12 months. Very high percentages of dystocia (40-55%) were found in the Mmp2-/- and dKO females, contrary to the Mmp9-/- and wild-type females. The histological analysis of the uterus and cervix revealed that Mmp2-/- tissues undergo marked structural alterations, including highly enlarged myometrial, endometrial and luminal cavity. Increased collagen deposition was also demonstrated, suggesting a mechanism of extensive fibrosis in the Mmp2-/- myometrium, which may result in dystocia. Overall, this study describes a new role for MMP2 in myometrium remodeling during mammalian parturition process, highlighting a novel cause for dystocia due to a loss in MMP2 activity in the uterine tissue.

Effect of milk fat globules on growth and metabolism in rats fed an unbalanced diet.

We assessed the effects of supplementing milk fat globules (MFG) on the growth and development of the skeleton in rats fed a Western unbalanced diet (UBD). The UBD is high in sugar and fat, low in protein, fiber, and micronutrients, and negatively impacts health. The MFG-a complex lipid-protein assembly secreted into milk-has a unique structure and composition, which differs significantly from isolated and processed dietary ingredients. Rats consuming the UBD exhibited growth retardation and disrupted bone structural and mechanical parameters; these were improved by supplementation with small MFG. The addition of small MFG increased the efficiency of protein utilization for growth, and improved trabecular and cortical bone parameters. Furthermore, consumption of UBD led to a decreased concentration of saturated fatty acids and increased levels of polyunsaturated fatty acids (PUFA), particularly omega-6 PUFA, in the serum, liver, and adipose tissue. The addition of small MFG restored PUFA concentration and the ratio of omega-6 to omega-3 PUFA in bone marrow and adipose tissue. Finally, large but not small MFG supplementation affected the cecal microbiome in rats. Overall, our results suggest that natural structure MFG supplementation can improve metabolism and bone development in rats fed an UBD, with the effects depending on MFG size. Moreover, the benefits of small MFG to bone development and metabolism were not mediated by the microbiome, as the detrimental effects of an UBD on the microbiome were not mitigated by MFG supplementation.

The Use of Post-Natal Skeleton Development as Sensitive Preclinical Model to Test the Quality of Alternative Protein Sources in the Diet.

Dietary protein is necessary throughout all life stages. Adequate intake of protein during juvenile years is essential to enable appropriate synthesis of bone matrix and achieve the full peak bone mass (PBM). Due to socio-demographic changes, accompanied by environmental damage and ethical problems, a transition to the consumption of different and alternative protein sources in the human diet must occur. This transition requires the precise evaluation of protein quality. Here, we utilize a preclinical model of young rats during their post-natal developmental period to define the nutritive quality of a number of alternative protein sources (soy, spirulina, chickpea, and fly larvae) by their health impact on growth performance and skeletal development. We indicate that when restricted (10% of calories) not one of the tested alternative protein sources have succeeded in causing optimal growth, as compared to the referenced source, casein; yet fly larvae protein followed by chickpea flour were found to be superior to the rest. Growth-plate histology and µ-CT analyses demonstrated a number of changes in growth patterns and bone morphometric parameters. Bone mechanical testing, by three-point bending analyses, was sensitive in demonstrating the effect of the reduction in the amount of the dietary protein. Moreover, the rats' weight and length, as well as their eating patterns, were found to reflect the proteins' quality better than their amino acid composition. Hence, our study emphasizes the importance of evaluating protein as a whole food source, and suggests a new approach for this purpose.

Nutritional Approaches as a Treatment for Impaired Bone Growth and Quality Following the Consumption of Ultra-Processed Food.

The severe impairment of bone development and quality was recently described as a new target for unbalanced ultra-processed food (UPF). Here, we describe nutritional approaches to repair this skeletal impairment in rats: supplementation with micro-nutrients and a rescue approach and switching the UPF to balanced nutrition during the growth period. The positive effect of supplementation with multi-vitamins and minerals on bone growth and quality was followed by the formation of mineral deposits on the rats' kidneys and modifications in the expression of genes involved in inflammation and vitamin-D metabolism, demonstrating the cost of supplementation. Short and prolonged rescue improved trabecular parameters but incompletely improved the cortical parameters and the mechanical performance of the femur. Cortical porosity and cartilaginous lesions in the growth-plate were still detected one week after rescue and were reduced to normal levels 3 weeks after rescue. These findings highlight bone as a target for the effect of UPF and emphasize the importance of a balanced diet, especially during growth.

The Use of Mushrooms and Spirulina Algae as Supplements to Prevent Growth Inhibition in a Pre-Clinical Model for an Unbalanced Diet.

Today's eating patterns are characterized by the consumption of unbalanced diets (UBDs) resulting in a variety of health consequences on the one hand, and the consumption of dietary supplements in order to achieve overall health and wellness on the other. Balanced nutrition is especially crucial during childhood and adolescence as these time periods are characterized by rapid growth and development of the skeleton. We show the harmful effect of UBD on longitudinal bone growth, trabecular and cortical bone micro-architecture and bone mineral density; which were analyzed by micro-CT scanning. Three point bending tests demonstrate the negative effect of the diet on the mechanical properties of the bone material as well. Addition of Spirulina algae or Pleurotus eryngii or Agaricus bisporus mushrooms, to the UBD, was able to improve growth and impaired properties of the bone. 16SrRNA Sequencing identified dysbiosis in the UBD rats' microbiota, with high levels of pro-inflammatory associated bacteria and low levels of bacteria associated with fermentation processes and bone related mechanisms. These results provide insight into the connection between diet, the skeletal system and the gut microbiota, and reveal the positive impact of three chosen dietary supplements on bone development and quality presumably through the microbiome composition.

Ultra-processed food targets bone quality via endochondral ossification.

Ultra-processed foods have known negative implications for health; however, their effect on skeletal development has never been explored. Here, we show that young rats fed ultra-processed food rich in fat and sugar suffer from growth retardation due to lesions in their tibial growth plates. The bone mineral density decreases significantly, and the structural parameters of the bone deteriorate, presenting a sieve-like appearance in the cortices and poor trabecular parameters in long bones and vertebrae. This results in inferior mechanical performance of the entire bone with a high fracture risk. RNA sequence analysis of the growth plates demonstrated an imbalance in extracellular matrix formation and degradation and impairment of proliferation, differentiation and mineralization processes. Our findings highlight, for the first time, the severe impact of consuming ultra-processed foods on the growing skeleton. This pathology extends far beyond that explained by the known metabolic effects, highlighting bone as a new target for studies of modern diets.

The Role of Omega-3 Polyunsaturated Fatty Acids from Different Sources in Bone Development.

N-3 polyunsaturated fatty acids (PUFAs) are essential nutrients that must be obtained from the diet. We have previously showed that endogenous n-3 PUFAs contribute to skeletal development and bone quality in fat-1 mice. Unlike other mammals, these transgenic mice, carry the n-3 desaturase gene and thus can convert n-6 to n-3 PUFAs endogenously. Since this model does not mimic dietary exposure to n-3 PUFAs, diets rich in fish and flaxseed oils were used to further elucidate the role of n-3 PUFAs in bone development. Our investigation reveals that dietary n-3 PUFAs decrease fat accumulation in the liver, lower serum fat levels, and alter fatty acid (FA) content in liver and serum. Bone analyses show that n-3 PUFAs improve mechanical properties, which were measured using a three-point bending test, but exert complex effects on bone structure that vary according to its source. In a micro-CT analysis, we found that the flaxseed oil diet improves trabecular bone micro-architecture, whereas the fish oil diet promotes higher bone mineral density (BMD) with no effect on trabecular bone. The transcriptome characterization of bone by RNA-seq identified regulatory mechanisms of n-3 PUFAs via modulation of the cell cycle and peripheral circadian rhythm genes. These results extend our knowledge and provide insights into the molecular mechanisms of bone remodeling regulation induced by different sources of dietary n-3 PUFAs.