SummaryMitochondrial reactive oxygen species (ROS) are proposed to play a central role in aging and ageassociated disorders, although direct in vivo evidence is lacking. We recently generated a mouse mutant with mutated Inner Mitochondrial Membrane Peptidase 2-like (Immp2l) gene, which impairs the signal peptide sequence processing of mitochondrial proteins cytochrome c1 and glycerol phosphate dehydrogenase 2. The mitochondria from mutant mice generate elevated levels of superoxide ion and cause impaired fertility in both sexes. Here we design experiments to examine the effects of excessive mitochondrial ROS generation on health span. We show that Immp2l mutation increases oxidative stress in multiple organs such as the brain and the kidney, although expression of superoxide dismutases in these tissues of the mutants is also increased. The mutants show multiple aging-associated phenotypes, including wasting, sarcopenia, loss of subcutaneous fat, kyphosis and ataxia, with female mutants showing earlier onset and more severe age-associated disorders than male mutants. The loss of body weight and fat was unrelated to food intake. Adipose derived stromal cells (ADSC) from mutant mice showed impaired proliferation capability, formed significantly less and smaller colonies in colony formation assays, although they retained adipogenic differentiation capability in vitro. This functional impairment was accompanied by increased levels of oxidative stress. Our data showed that mitochondrial ROS is the driving force of accelerated aging and suggested that ROS damage to adult stem cells could be one of the mechanisms for age-associated disorders.

Human hematopoietic niches are complex specialized microenvironments that maintain and regulate hematopoietic stem and progenitor cells (HSPC). Thus far, most of the studies performed investigating alterations of HSPCniche dynamic interactions are conducted in animal models. Herein, organ microengineering with microfluidics is combined to develop a human bone marrow (BM)-on-a-chip with an integrated recirculating perfusion system that consolidates a variety of important parameters such as 3D architecture, cell-cell/cell-matrix interactions, and circulation, allowing a better mimicry of in vivo conditions. The complex BM environment is deconvoluted to 4 major distinct, but integrated, tissue-engineered 3D niche constructs housed within a single, closed, recirculating microfluidic device system, and equipped with cell tracking technology. It is shown that this technology successfully enables the identification and quantification of preferential interactions-homing and retention-of circulating normal and malignant HSPC with distinct niches.

Mitochondrial reactive oxygen species (ROS) have been implicated in spermatogenic damage, although direct in vivo evidence is lacking. We recently generated a mouse in which the Inner Mitochondrial Membrane Peptidase 2-like (Immp2l) gene is mutated. This Immp2l mutation impairs the processing of signal peptide sequences from mitochondrial cytochrome c1 and glycerol phosphate dehydrogenase 2. The mitochondria from mutant mice generate elevated levels of superoxide ion, which causes age-dependent spermatogenic damage. Here we confirm age-dependent spermatogenic damage in a new cohort of mutants, which started at the age of 10.5 months. Compared with age-matched controls, protein carbonyl content was normal in testes of 2- to 5-month-old mutants, but significantly elevated in testes of 13-month-old mutants, indicating elevated oxidative stress in the testes at the time of impaired spermatogenesis. Testicular expression of superoxide dismutases was not different between control and mutant mice, while that of catalase was increased in young and old mutants. The expression of cytosolic glutathione peroxidase 4 (phospholipid hydroperoxidase) in testes was significantly reduced in 13-month-old mutants, concomitant with impaired spermatogenesis. Apoptosis of all testicular populations was increased in mutant mice with spermatogenic damage. The mitochondrial DNA (mtDNA) mutation rate in germ cells of mutant mice with impaired spermatogenesis was unchanged, excluding a major role of mtDNA mutation in ROS-mediated spermatogenic damage. Our data show that increased mitochondrial ROS are one of the driving forces for spermatogenic impairment.

bThe function of MEX3C, the mammalian homolog of Caenorhabditis elegans RNA-binding protein muscle excess 3 (MEX-3), was unknown until our recent report that MEX3C is necessary for normal postnatal growth and enhances the expression of local bone Igf1 expression. Here we report the pivotal role of Mex3c in energy balance regulation. Mex3c mutation caused leanness in both heterozygous and homozygous transgenic mice, as well as a more beneficial blood glucose and lipid profile in homozygous transgenic mice, in both sexes. Although transgenic mice showed normal food intake and fecal lipid excretion, they had increased energy expenditure independent of physical activity. Mutant mice had normal body temperature, Ucp1 expression in brown adipose tissue, and muscle and liver fatty acid oxidation. Mex3c is expressed in neurons and is detectable in the arcuate nucleus, the ventromedial nucleus, and the dorsomedial nucleus of the hypothalamus. Mex3c was not detected in NPY or POMC neurons but was detected in leptin-responsive neurons in the ventromedial nucleus. Mex3c and Leptin double mutant mice were growth retarded and obese and had blood profiles similar to those of ob/ob mice but showed none of the steatosis observed in ob/ob mice. Our data show that Mex3c is involved in energy balance regulation. Caenorhabditis elegans MEX-3 is an hnRNP K homology (KH) domain-containing RNA-binding protein regulating RNA targets such as pal-1 (13, 22, 23), rme-2 (6), and nos-2 (25). It is involved in the cell fate specification process in C. elegans and in totipotency maintenance of the germ line in adult worms (7,13,23,40). Human and mouse genomes encode four MEX-3 homologues: MEX3A, MEX3B, MEX3C, and MEX3D (3). They all have two KH RNA-binding domains at the N terminus which are also present in the C. elegans homolog and a zinc finger (ZNF) domain at the C terminus which is absent in MEX-3 in C. elegans. MEX3A and MEX3B colocalize with decapping factor DCP1a and Argonaute proteins in processing bodies (3), and the localization of MEX3B is regulated by 14-3-3 protein (9). MEX3D, once called TINO, is a BCL2 mRNA AU-rich element-binding protein that negatively regulates the stability of BCL2 mRNA (12).Mouse MEX3C is 99% identical to human, chimpanzee, and bovine MEX3Cs. Linkage analysis and association studies suggest that MEX3C contributes to genetic susceptibility of hypertension (21), although the mechanism is unknown. Recently, we reported that Mex3c mutation in mice causes growth retardation due to IGF1 deficiency in developing bone (26). Mex3c is highly expressed in resting and proliferating chondrocytes, and IGF1 protein expression in these cells of mutant mice is significantly reduced, although Igf1 mRNA expression in bones from mutant mice is not changed. In the C57BL/6 background, 85% of homozygous mutant pups die soon after birth, whereas in the FVB/N background, about 80% of the pups survive to adulthood, although they are still growth retarded. Mutant male and female mice are fertile, although Mex3c is highly expressed i...

Renal disease is a worldwide health issue. Besides transplantation, current therapies revolve around dialysis, which only delays disease progression but cannot replace other renal functions, such as synthesizing erythropoietin. To address these limitations, cell‐based approaches have been proposed to restore damaged kidneys as an alternative to current therapies. Recent studies have shown that stem cell‐derived secretomes can enhance tissue regeneration. However, many growth factors undergo rapid degradation when they are injected into the body in a soluble form. Efficient delivery and controlled release of secreting factors at the sites of injury would improve the efficacy in tissue regeneration. Herein, we developed a gel‐based delivery system for controlled delivery of trophic factors in the conditioned medium (CM) secreted from human placental stem cells (HPSCs) and evaluated the effect of trophic factors on renal regeneration. CM treatment significantly enhanced cell proliferation and survival in vitro. Platelet‐rich plasma (PRP) was used as a delivery vehicle for CM. Analysis of the release kinetics demonstrated that CM delivery through the PRP gel resulted in a controlled release of the factors both in vitro and in vivo. In an acute kidney injury model in rats, functional and structural analysis showed that CM delivery using the PRP gel system into the injured kidney minimized renal tissue damage, leading to a more rapid functional recovery when compared with saline, CM, or vehicle only injection groups. These results suggest that controlled delivery of HPSC‐derived trophic factors may provide efficient repair of renal tissue injury. stem cells translational medicine 2019;8:959&970

Improved methods of cell culture from eye stalk, hepatopancreas, muscle, ovary, and hemocytes of shrimp (Penaeus vannamei) were established using synthetic media and shrimp muscle extract (SME). For hemocytes and ovarian cell cultures, Grace's insect medium supplemented with 10% (v/v) fetal bovine serum and 10% SME (v/v) showed enhanced attachment and proliferation of the cells. The hemocyte and ovarian cell cultures could be maintained for 48 and 66 days, respectively, and have been sub-cultured four and six times, respectively. Both ovary and hemocyte cell cultures contained primarily epithelial-like cells. Cells derived from ovary tissue grew preferably between 26°C and 28°C with 5% CO(2). Although the temperature preference of hemocyte cells was the same as ovarian cells, CO(2) supplementation did not show any difference in the growth of hemocyte cells. When the shrimp were injected with lipopolysaccharide (8 μg/g of shrimp) and hemolymph was drawn 24 h post-injection, the in vitro multiplicity of hemocytes dramatically improved. The growth of eye stalk, hepatopancreas, and muscle-derived cells was much less compared to ovarian cells and hemocytes under the conditions described above. The optimal culture conditions for ovarian cells and hemocytes were also different from that for eye stalk, hepatopancreas, and muscle cell culture. The proliferation efficiencies of primary cultures of hepatopancreas, eyestalk, and muscle cells were about 30, 12, and <7 d, respectively. The improved culture conditions described here, particularly for hemocytes and ovary, will be very useful for in vitro studies involving viruses infecting shrimp and in shrimp genomic studies.

Tracheal stenosis is a rare but life-threatening disease. Primary clinical procedures for treating this disease are limited if the region requiring repair is long or complex. This study is the first of its kind to fabricate bioprinted tracheal constructs with separate cartilage and smooth muscle regions using polycaprolactone (PCL) and human mesenchymal stem cell (hMSC)-laden hydrogels. Our final bioprinted trachea showed comparable elastic modulus and yield stress compared to native tracheal tissue. In addition, both cartilage and smooth muscle formation were observed in the desired regions of our bioprinted trachea through immunohistochemistry and western blot after two weeks of in vitro culture. This study demonstrates a novel approach to manufacture tissue engineered trachea with mechanical and biological properties similar to native trachea, which represents a step closer to overcoming the clinical challenges of treating tracheal stenosis.

Future deep space missions to Mars and near-Earth asteroids will expose astronauts to chronic solar energetic particles (SEP) and galactic cosmic ray (GCR) radiation, and likely one or more solar particle events (SPEs). Given the inherent radiosensitivity of hematopoietic cells and short latency period of leukemias, space radiation-induced hematopoietic damage poses a particular threat to astronauts on extended missions. We show that exposing human hematopoietic stem/progenitor cells (HSC) to extended mission-relevant doses of accelerated high-energy protons and iron ions leads to the following: (1) introduces mutations that are frequently located within genes involved in hematopoiesis and are distinct from those induced by γ-radiation; (2) markedly reduces in vitro colony formation; (3) markedly alters engraftment and lineage commitment in vivo; and (4) leads to the development, in vivo, of what appears to be T-ALL. Sequential exposure to protons and iron ions (as typically occurs in deep space) proved far more deleterious to HSC genome integrity and function than either particle species alone. Our results represent a critical step for more accurately estimating risks to the human hematopoietic system from space radiation, identifying and better defining molecular mechanisms by which space radiation impairs hematopoiesis and induces leukemogenesis, as well as for developing appropriately targeted countermeasures.