A NEUROGENIC GENE EXPRESSION SIGNATURE SUPPORTS HUMAN THERMOGENIC ADIPOSE TISSUE DEVELOPMENT IN VIVO.

Human beige/brite thermogenic adipose tissue exerts beneficial metabolic effects and may be harnessed to improve metabolic health. To uncover mechanisms by which thermogenic adipose tissue is generated and maintained we developed a species-hybrid model in which human mesenchymal progenitor cells are induced in vitro to differentiate into white or thermogenic adipocytes and are then implanted into immuno-compromised mice. Upon implantation, thermogenic adipocytes form a more densely vascularized and innervated adipose tissue compared to non-thermogenic adipocytes. Mouse endothelial and stem/progenitor cells recruited by implanted human thermogenic adipocytes are also qualitatively different, with differentially expressed genes mapping predominantly to circadian rhythm pathways. We trace the formation of this enhanced neurovascular architecture to higher expression of a distinct set of genes directly associated with neurogenesis (THBS4, TNC, NTRK3 and SPARCL1), and to lower expression of genes associated with neurotransmitter degradation (MAOA, ACHE) by adipocytes in the developed tissue. Further analysis reveals that MAOA is abundant in human adipocytes but absent in mouse adipocytes, revealing species-specific mechanisms of neurotransmitter tone regulation. In summary, our work discovers specific neurogenic genes associated with development and maintenance of human thermogenic adipose tissue, reveals species-specific mechanisms of control of neurotransmitter tone, and suggests that targeting adipocyte MAOA may be a strategy for enhancing thermogenic adipose tissue activity in humans.

3D Texture Analysis of MRI Relaxation Time Maps for Assessment of Repair Cartilage with Treatment of Allogeneic Human Adipose-Derived Mesenchymal Progenitor Cells

Background: We used textural analysis matrix to examine the spatial distribution of pixel values and detect the compositional variation of repair cartilage with treatment of allogeneic human adipose-derived mesenchymal progenitor cells (haMPCs). Methods: Eighteen patients were divided randomly into three groups with intra-articular injections of haMPCs: the low-dose (1.0×107 cells), mid-dose (2.0×107), and high-dose (5.0×107) group with six patients each. 3D texture analyses based on gray level run-length matrix (GLRLM) of the segmented ROIs on MRI relaxation time maps including T1rho, T2, T2* and R2*. Five GLRLM parameters were analyzed, including run length non-uniformity (RLNonUni), grey level non-uniformity (GLevNonU), long run emphasis (LngREmph), short run emphasis (ShrtREmp) and fraction of image in runs (Fraction). We used the difference before and after treatment (D values) as the object to avoid errors caused by individual differences. Two-tailed Pearson linear correlation analysis was used to investigate correlations between texture parameters and the WOMAC scores. Results: The heterogeneity of spatial distribution of MRI relaxation time mapping pixels from three groups was decreased to varying degrees at 48 weeks after intra-articular injection of haMPCs. Spatial distribution of cartilage relaxation time maps pixels were uneven and layered, especially in T2 maps. Compared with base time, there were significant differences among three dose groups in GLRLM features for T1rho map including RLNonUni, GLevNonU, LngREmph, for T2 map including LngREmph, GLevNonU, ShrtREmp, for T2* map including RLNonUni, GLevNonU, and for R2* map including RLNonUni, GLevNonU. WOMAC pain scores were associated with RLNonUni of T1rho map, GLevNonU of T2 map, LngREmph of T2* map, LngREmph of R2* map and Fraction of T1rho map, whereas no significant correlations in other measurements.Conclusions: MRI texture analysis of cartilage may allow detection of the compositional variation of repair cartilage with treatment of allogeneic haMPCs. This has potential applications in understanding mechanism of stem cells repairing cartilage and assessing response to treatment.Trial registration: Clinicaltrials, NCT02641860. Registered 3 December 2015.https://www.clinicaltrials.gov/ct2/show/NCT02641860

How Does Testosterone Augment the Anabolic Response to Exercise in Frail Older Adults?

Testosterone treatment increases muscle mass, strength, and leg power in menopausal women, hypogonadal men, older men with mobility limitation, COPD and ESRD. Testosterone's effects on muscle mass and strength are augmented by exercise training and growth hormone. Testosterone treatment improves some measures of physical performance, such as stair climbing power and aerobic capacity; the improvements in gait speed have been modest. Testosterone increases muscle mass by inducing the hypertrophy of type 1 and 2 muscle fibers, and by increasing satellite cell number. Testosterone promotes the differentiation of mesenchymal progenitor cells into myogenic lineage and inhibits their differentiation into adipogenic lineage by activating Wnt-target genes, including follistatin that plays an important role in mediating testosterone's effects on the muscle. Testosterone also increases polyamine synthesis in the muscle. Combined administration of testosterone plus multi-component exercise intervention that includes functional training may be needed to improve function and mobility in older adults.

Gene-Activated Matrix with Self-Assembly Anionic Nano-Device Containing Plasmid DNAs for Rat Cranial Bone Augmentation

We have developed nanoballs, a biocompatible self-assembly nano-vector based on electrostatic interactions that arrange anionic macromolecules to polymeric nanomaterials to create nucleic acid carriers. Nanoballs exhibit low cytotoxicity and high transfection efficiently in vivo. This study investigated whether a gene-activated matrix (GAM) composed of nanoballs containing plasmid (p) DNAs encoding bone morphogenetic protein 4 (pBMP4) could promote bone augmentation with a small amount of DNA compared to that composed of naked pDNAs. We prepared nanoballs (BMP4-nanoballs) constructed with pBMP4 and dendrigraft poly-L-lysine (DGL, a cationic polymer) coated by γ-polyglutamic acid (γ-PGA; an anionic polymer), and determined their biological functions in vitro and in vivo. Next, GAMs were manufactured by mixing nanoballs with 2% atelocollagen and β-tricalcium phosphate (β-TCP) granules and lyophilizing them for bone augmentation. The GAMs were then transplanted to rat cranial bone surfaces under the periosteum. From the initial stage, infiltrated macrophages and mesenchymal progenitor cells took up the nanoballs, and their anti-inflammatory and osteoblastic differentiations were promoted over time. Subsequently, bone augmentation was clearly recognized for up to 8 weeks in transplanted GAMs containing BMP4-nanoballs. Notably, only 1 μg of BMP4-nanoballs induced a sufficient volume of new bone, while 1000 μg of naked pDNAs were required to induce the same level of bone augmentation. These data suggest that applying this anionic vector to the appropriate matrices can facilitate GAM-based bone engineering.

The Treasury of Wharton's Jelly

Background Postnatal umbilical cord tissue contains valuable mesenchymal progenitor cells of various differentiation stages. While mesenchymal stem cells are plastic-adherent and tend to differentiate into myofibroblastic phenotypes, some round cells detach, float above the adherent cells, and build up cell aggregates, or form spheroids spontaneously. Very small luminescent cells are always involved as single cells or within collective forms and resemble the common well-known very small embryonic-like cells (VSELs). In this study, we investigated these VSELs-like cells in terms of their pluripotency phenotype and tri-lineage differentiation potential. Methods VSELs-like cells were isolated from cell-culture supernatants by a process that combines filtering, up concentration, and centrifugation. To determine their pluripotency character, we measured the expression of Nanog, Sox-2, Oct-4, SSEA-1, CXCR4, SSEA-4 on gene and protein level. In addition, the cultured cells derived from UC tissue were examined regarding their potential to differentiate into three germ layers. Result The VSELs-like cells express all of the pluripotency-associated markers we investigated and are able to differentiate into meso- endo- and ectodermal precursor cells. Conclusions Umbilical cord tissue hosts highly potent VSELs-like stem cells. Graphical Abstract

Irisin Attenuates Osteogenic Differentiation of Mouse Mesenchymal Stem Cells via Suppressing Bone Morphogenetic Protein-2 Signaling

This study assessed the impact of irisin on mouse mesenchymal stem cells (MSCs) differentiation and the underlying mechanisms. Bone marrow mesenchymal stem cells (BMMSCs) and mesenchymal progenitor cells (KUSA-A1 cells) were isolated from mice and inoculated into petri dishes. Cell proliferation and apoptosis under different concentrations of irisin were detected by MTT. Irisin (1 μM) with nontoxic dose concentration was selected for subsequent experiments. Cells were exposed to 1 μM irisin, osteogenic differentiation was detected by von kossa stain, we employed oil red o stain to test adipocyte differentiation, Alcian blue stain to determine chondrocyte differentiation. BMP-2 expression was analyzed by immunocytochemical staining. Finally, signal transduction pathways and expression and transcription levels of osteogenic differentiation markers in irisin-treated cells were detected by protein imprinting and PCR. BMMSCs and KUSA-A1 cells displayed significantly suppressed osteogenic differentiation and reduced formation of extracellular mineralized matrix, while BMMSCs presented unaffected adipocyte differentiation and chondrocyte differentiation. 1 μM Irisin did not exert cytotoxicity. Further, irisin treatment abated osteogenic differentiation makers Runx2, Osterix, Osteocalcin, Osteopontin, alkaline phosphatase expression. Finally, BMP-2/Smads signaling related molecules (BMP-2, Smad1, Smad4, Smad5 and Smad8) levels were reduced after Irisin treatment. Irisin might inhibit osteoblast differentiation by modulating BMP-2/Smads axis.

Highly Segregated Biocomposite Membrane as a Functionally Graded Template for Periodontal Tissue Regeneration

Guided tissue regeneration (GTR) membranes are used for treating chronic periodontal lesions with the aim of regenerating lost periodontal attachment. Spatially designed functionally graded bioactive membranes with surface core layers have been proposed as the next generation of GTR membranes. Composite formulations of biopolymer and bioceramic have the potential to meet these criteria. Chitosan has emerged as a well-known biopolymer for use in tissue engineering applications due to its properties of degradation, cytotoxicity and antimicrobial nature. Hydroxyapatite is an essential component of the mineral phase of bone. This study developed a GTR membrane with an ideal chitosan to hydroxyapatite ratio with adequate molecular weight. Membranes were fabricated using solvent casting with low and medium molecular weights of chitosan. They were rigorously characterised with scanning electron microscopy, Fourier transform infrared spectroscopy in conjunction with photoacoustic sampling accessory (FTIR-PAS), swelling ratio, degradation profile, mechanical tensile testing and cytotoxicity using human osteosarcoma and mesenchymal progenitor cells. Scanning electron microscopy showed two different features with 70% HA at the bottom surface packed tightly together, with high distinction of CH from HA. FTIR showed distinct chitosan dominance on top and hydroxyapatite on the bottom surface. Membranes with medium molecular weight showed higher swelling and longer degradation profile as compared to low molecular weight. Cytotoxicity results indicated that the low molecular weight membrane with 30% chitosan and 70% hydroxyapatite showed higher viability with time. Results suggest that this highly segregated bilayer membrane shows promising potential to be adapted as a surface layer whilst constructing a functionally graded GTR membrane on its own and for other biomedical applications.

Rapid Production and Genetic Stability of Human Mesenchymal Progenitor Cells Derived from Human Somatic Cell Nuclear Transfer-Derived Pluripotent Stem Cells

Pluripotent stem cell-derived mesenchymal progenitor cells (PSC-MPCs) are primarily derived through two main methods: three-dimensional (3D) embryoid body-platform (EB formation) and the 2D direct differentiation method. We recently established somatic cell nuclear transfer (SCNT)-PSC lines and showed their stemness. In the present study, we produced SCNT-PSC-MPCs using a novel direct differentiation method, and the characteristics, gene expression, and genetic stability of these MPCs were compared with those derived through EB formation. The recovery and purification of SCNT-PSC-Direct-MPCs were significantly accelerated compared to those of the SCNT-PSC-EB-MPCs, but both types of MPCs expressed typical surface markers and exhibited similar proliferation and differentiation potentials. Additionally, the analysis of gene expression patterns using microarrays showed very similar patterns. Moreover, array CGH analysis showed that both SCNT-PSC-Direct-MPCs and SCNT-PSC-EB-MPCs exhibited no significant differences in copy number variation (CNV) or single-nucleotide polymorphism (SNP) frequency. These results indicate that SCNT-PSC-Direct-MPCs exhibited high genetic stability even after rapid differentiation into MPCs, and the rate at which directly derived MPCs reached a sufficient number was higher than that of MPCs derived through the EB method. Therefore, we suggest that the direct method of differentiating MPCs from SCNT-PSCs can improve the efficacy of SCNT-PSCs applied to allogeneic transplantation.

MicroRNA-29a in Osteoblasts Represses High-Fat Diet-Mediated Osteoporosis and Body Adiposis through Targeting Leptin

Skeletal tissue involves systemic adipose tissue metabolism and energy expenditure. MicroRNA signaling controls high-fat diet (HFD)-induced bone and fat homeostasis dysregulation remains uncertain. This study revealed that transgenic overexpression of miR-29a under control of osteocalcin promoter in osteoblasts (miR-29aTg) attenuated HFD-mediated body overweight, hyperglycemia, and hypercholesterolemia. HFD-fed miR-29aTg mice showed less bone mass loss, fatty marrow, and visceral fat mass together with increased subscapular brown fat mass than HFD-fed wild-type mice. HFD-induced O2 underconsumption, respiratory quotient repression, and heat underproduction were attenuated in miR-29aTg mice. In vitro, miR-29a overexpression repressed transcriptomic landscapes of the adipocytokine signaling pathway, fatty acid metabolism, and lipid transport, etc., of bone marrow mesenchymal progenitor cells. Forced miR-29a expression promoted osteogenic differentiation but inhibited adipocyte formation. miR-29a signaling promoted brown/beige adipocyte markers Ucp-1, Pgc-1α, P2rx5, and Pat2 expression and inhibited white adipocyte markers Tcf21 and Hoxc9 expression. The microRNA also reduced peroxisome formation and leptin expression during adipocyte formation and downregulated HFD-induced leptin expression in bone tissue. Taken together, miR-29a controlled leptin signaling and brown/beige adipocyte formation of osteogenic progenitor cells to preserve bone anabolism, which reversed HFD-induced energy underutilization and visceral fat overproduction. This study sheds light on a new molecular mechanism by which bone integrity counteracts HFD-induced whole-body fat overproduction.