Intrathecal delivery of human ESC-derived mesenchymal stem cell spheres promotes recovery of a primate multiple sclerosis model.

Nonhuman primate experimental autoimmune encephalomyelitis (EAE) is a valuable model for multiple sclerosis, an inflammatory demyelinating disease in the central nervous system (CNS). Human embryonic stem cell-derived mesenchymal stem cells (EMSC) are effective in treating murine EAE. Yet, it remains unknown whether the EMSC efficacy is translatable to humans. Here we induced a primate EAE model in cynomolgus monkeys and delivered EMSC in spheres (EMSCsp) to preserve the cell viability during long-distance transportation. EMSCsp intrathecally injected into the CNS, remarkably reduced the clinical symptoms, brain lesions, and neuronal demyelination in the EAE monkeys during a 3-month observation. Whereas, symptoms in the vehicle control-injected EAE monkey remained and reduced slowly and MRI lesions in brain expanded. Moreover, EMSC could transdifferentiate into neural cells in vivo in the CNS of the treated animals. Supporting evidence demonstrated that EMSCsp cells cultured in cerebrospinal fluid from the EAE monkeys largely converted to neural cells with elevated expression of genes for neuronal markers, neurotrophic factors, and neuronal myelination. Thus, this study demonstrates that EMSCsp injected directly into the CNS, can attenuate the disease progression in the primate EAE model, highly encouraging for clinical translation.

Cryopreservation of cynomolgus macaque (Macaca fascicularis) sperm with glycerol and ethylene glycol, and its effect on sperm-specific ion channels - CatSper and Hv1.

The cryoprotective agent (CPA) is one of the most important factors that affects the cryosurvival of sperm. The aim of the present study was to compare two different CPAs, glycerol (Gly) and ethylene glycol (EG), on the cryopreservation of cynomolgus macaques sperm and evaluate the effects of cryopreservation on sperm motility, acrosomal integrity, DNA integrity, mitochondrial function and the sperm membrane ion channels CatSper and Hv1. Compared to fresh sperm, cryopreservation with either 0.7 M Gly or EG decreased the sperm motility (79.8 ± 1.5% Vs. 47.3 ± 1.8% and 47.6 ± 1.4%), acrosomal integrity (89.6 ± 1.2% Vs. 80.1 ± 1.8% and 79.6 ± 1.7%), DNA integrity (91.9 ± 0.7% Vs. 82.9 ± 1.0% and 82.3 ± 1.0%) and mitochondrial membrane potential (87.9 ± 1.8% Vs. 70.6 ± 2.7% and 67.9 ± 2.5%) and the quantity of the CatSper and Hv1 channels determined by Western Blot (p < 0.05), and EG showed equal cryoprotection to cynomolgus sperm in all of the sperm parameters. Our results indicated, for the first time, that cryopreservation decreases the quantity of sperm membrane ion channels (CatSper and Hv1), which might be one of the reasons that frozen sperm have a low fertilizing ability. The study will be beneficial to understand the biological process involved in sperm cryopreservation of nonhuman primates and contribute to improving cryopreservation protocols than can maintain sperm function and fertilizing ability.

Cryopreservation of Cynomolgus Macaque (Macaca fascicularis) Sperm by Using a Commercial Egg-YolkFree Freezing Medium.

Conventional TRISegg yolk (TEY) freezing medium for the cryopreservation of NHP sperm has the risk of contamination due to widespread zoonotic diseases. This study was aimed at determining the optimal glycerol concentration, freezing rate, and holding time in liquid N2 vapor for the cryopreservation of cynomolgus macaque sperm by using a commercial egg-yolkfree freezing medium (SC medium) designed for human sperm cryopreservation. Sperm motility and acrosomal integrity after freezing were assessed. Sperm in SC medium (dilution ratio, 3:1) frozen at cooling rates of 67 and 183C/min in liquid N2 vapor showed higher post-thaw motility than did samples frozen at 435C/min. At the cooling rate of 183C/min and dilution in SC medium at a 3:1 ratio, post-thaw motility was higher after a holding time of 10 min than after 30 min (recommended by the manufacturer). In addition, post-thaw motility of sperm frozen in SC medium was higher with dilution ratios of 3:1, 4.5:1, and 6:1 compared with 9:1, 10.5:1, and 12:1, and the sample diluted 12:1 showed the lowest percentage of thawed sperm with intact acrosomes. Sperm showed higher post-thaw motility after freezing in TEY than in SC medium; acrosomal integrity did not differ between the 2 media. Our results indicated that cynomolgus macaque sperm can be cryopreserved successfully by using a commercial egg-yolkfree freezing medium, which provides an option for genetic preservation with decreased zoonotic risk in this important NHP species.

Rhesus monkey model of liver disease reflecting clinical disease progression and hepatic gene expression analysis.

Alcoholic liver disease (ALD) is a significant public health issue with heavy medical and economic burdens. The aetiology of ALD is not yet completely understood. The development of drugs and therapies for ALD is hampered by a lack of suitable animal models that replicate both the histological and metabolic features of human ALD. Here, we characterize a rhesus monkey model of alcohol-induced liver steatosis and hepatic fibrosis that is compatible with the clinical progression of the biochemistry and pathology in humans with ALD. Microarray analysis of hepatic gene expression was conducted to identify potential molecular signatures of ALD progression. The up-regulation of expression of hepatic genes related to liver steatosis (CPT1A, FASN, LEPR, RXRA, IGFBP1, PPARGC1A and SLC2A4) was detected in our rhesus model, as was the down-regulation of such genes (CYP7A1, HMGCR, GCK and PNPLA3) and the up-regulation of expression of hepatic genes related to liver cancer (E2F1, OPCML, FZD7, IGFBP1 and LEF1). Our results demonstrate that this ALD model reflects the clinical disease progression and hepatic gene expression observed in humans. These findings will be useful for increasing the understanding of ALD pathogenesis and will benefit the development of new therapeutic procedures and pharmacological reagents for treating ALD.