Exosomes derived from human umbilical cord mesenchymal stem cells alleviate Parkinson's disease and neuronal damage through inhibition of microglia.

Microglia-mediated inflammatory responses have been shown to play a crucial role in Parkinson's disease. In addition, exosomes derived from mesenchymal stem cells have shown anti-inflammatory effects in the treatment of a variety of diseases. However, whether they can protect neurons in Parkinson's disease by inhibiting microglia-mediated inflammatory responses is not yet known. In this study, exosomes were isolated from human umbilical cord mesenchymal stem cells and injected into a 6-hydroxydopamine-induced rat model of Parkinson's disease. We found that the exosomes injected through the tail vein and lateral ventricle were absorbed by dopaminergic neurons and microglia on the affected side of the brain, where they repaired nigral-striatal dopamine system damage and inhibited microglial activation. Furthermore, in an in vitro cell model, pretreating lipopolysaccharide-stimulated BV2 cells with exosomes reduced interleukin-1ß and interleukin-18 secretion, prevented the adoption of pyroptosis-associated morphology by BV2 cells, and increased the survival rate of SH-SY5Y cells. Potential targets for treatment with human umbilical cord mesenchymal stem cells and exosomes were further identified by high-throughput microRNA sequencing and protein spectrum sequencing. Our findings suggest that human umbilical cord mesenchymal stem cells and exosomes are a potential treatment for Parkinson's disease, and that their neuroprotective effects may be mediated by inhibition of excessive microglial proliferation.

Expression of recombinant human Apolipoprotein A-IMilano in Nicotiana tabacum.

Apolipoprotein A-IMilano (Apo A-IMilano) is a natural mutant of Apolipoprotein. It is currently the only protein that can clear arterial wall thrombus deposits and promptly alleviate acute myocardial ischemia. Apo A-IMilano is considered as the most promising therapeutic protein for treating atherosclerotic diseases without obvious toxic or side effects. However, the current biopharmaceutical platforms are not efficient for developing Apo A-IMilano. The objectives of this research were to express Apo A-IMilano using the genetic transformation ability of N. tabacum. The method is to clone the coding sequence of Apo A-IMilano into the plant binary expression vector pCHF3 with a Flag/His6/GFP tag. The constructed plasmid was transformed into N. tabacum by a modified agrobacterium-mediated method, and transformants were selected under antibiotic stress. PCR, RT-qPCR, western blot and co-localization analysis was used to further verify the resistant N. tabacum. The stable expression and transient expression of N. tabacum were established, and the pure product of Apo A-IMilano was obtained through protein A/G agarose. The results showed that Apo A-IMilano was expressed in N. tabacum with a yield of 0.05 mg/g leaf weight and the purity was 90.58% ± 1.65. The obtained Apo A-IMilano protein was subjected to amino acid sequencing. Compared with the theoretical sequence of Apo A-IMilano, the amino acid coverage was 86%, it is also found that Cysteine replaces Arginine at position 173, which indicates that Apo A-IMilano, a mutant of Apo A-I, is accurately expressed in N. tabacum. The purified Apo A-IMilano protein had a lipid binding activity. The established genetic modification N. tabacum will provide a cost-effective system for the production of Apo A-IMilano. Regarding the rapid propagation of N. tabacum, this system provides the possibility of large-scale production and accelerated clinical translation of Apo A-IMilano.

Efficient differentiation of neural stem cells induced by the rat bone marrow stromal cells.

Neural stem cells (NSCs) are valuable self-renewing cells that can maintain the capacity to differentiate into specific brain cell types. NSCs may repair and even replace the brain tissue, and ultimatley promoting the central nervous system regeneration. Therefore, it is important, for scientists and pjysicians, to study the method for efficient culture and differentiation of NSCs. Our previous study demonstrated that Bone Marrow Stromal Cells (BMSCs) can directly regulate the differentiation of NSCs into neurons, and soluble molecules excreted by BMSCs played a key role in this process. Hereby, we further identified the BMSCs-induced neurons could form the synapses, convey dopamine and express voltage-depend and receptor-depend calcium channels. Moreover, the extracellular signal-regulated protein kinase ERK1/2 pathway was founded to be involved in the process of neuron differentiation and proliferation by the in vitro experiments. Finally, by using protein array, we, for the first time, found that the cytokine-induced neutrophil chemoattractant-3 (CINC-3, a small molecule cytokine) can promote the leukocytes invasion into the inflammation site, and have the ability to induce mesencephal NSCs into neurons. Consequently, these positive findings suggested that our BMSCs-induced culture system could provide a useful tool to investigate the molecular mechanisms of neural differentiation of NSCs, which may be benifical for neurodegenerative diseases in the near future.

Microglial activation and age-related dopaminergic neurodegeneration in MPTP-treated SAMP8 mice.

Senescence-accelerated mouse prone 8 (SAMP8) has an early onset of senility and a shorter life span, providing with cognitive impairment. Contrasted with C57BL/6 mouse, which is most commonly used in the study of Parkinson's disease (PD), SAMP8 needs shorter period of breeding and might be good candidate for the investigation of cognitive impairment in PD. Studies had shown the increase of sensibility to 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) with aging in C57BL/6 mouse. However, the sensitivity of MPTP neurotoxicity depends on the strains of animal and the exact mechanisms of the progression of PD promoted by aging is lack of consensus. Here, we showed after MPTP injection, the spontaneous activity of both young (3-month-old) and old (6-month-old) SAMP8 decreased dramatically, and the old mice required longer recovery time. Immunohistochemical and immunoblot analysis revealed that old mice displayed significant reductions in the dopaminergic neuron numbers and tyrosine hydroxylase (TH) protein. Microglia protein (CD11b) in the striatum of old mice increased more pronouncedly than that in the young mice from 24 h to 3 days. Inducible nitric oxide synthase (iNOS) in the striatum remarkably increased, however, no discernible difference between the two groups was found. These results suggested that the sensibility to MPTP increased with aging in SAMP8. A greater increase of microglial activation in old mice may be a possible mechanism to explain how advancing age predisposes the dopamine system to parkinsonism. The MPTP-SAMP8 model will start a new consideration for the study of PD.

Neuroprotective effects of enriched environment in MPTP-treated SAMP8 mice.

Neuroprotective effects of enriched environment (EE) have been well established. Recent study suggests that exposure to EE can protect dopaminergic neurons against MPTP-induced Parkinsonism. After 64 female SAMP8 mice were reared in EE and standard environment (SE) for 3 months, the effects of EE and SE were compared on behavioural change, tyrosine hydroxylase (TH) immunoreaction positive neuron and dopaminetransporter (DAT) expression in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)-treated SAMP8. EE mice showed decreased spontaneous activity compared with SE mice. But EE+MPTP mice showed less decreased spontaneous activity compared with SE+MPTP mice. Otherwise, EE mice showed increased percentage of entries into the open arms and percentage of time spent in the open arms. Furthermore, EE mice demonstrated reduced neurotoxicity, with less decreased TH mRNA and protein expression in Substantia Nigra (SN) after MPTP administration compared with SE mice. SE mice showed a 53.77% loss of TH-positive neurons, whereas EE mice only showed a 42.28% loss. Moreover, EE mice showed decreased DAT mRNA and protein expression compared with SE mice. These data demonstrate that EE can protect dopaminergic neurons against MPTP-induced neuronal damage, which suggest that the probability of developing Parkinson's disease (PD) may be related to life environment.

[Circadian rhythm of melatonin and blood pressure changes in patients with essential hypertension].

OBJECTIVE: To observe the association between melatonin (MLT) secretion and blood pressure changes during 24 hours in hypertensive patients (HPT) with (dipping) or without (non-dipping) night time blood pressure reduction. METHODS: The 24-hour blood pressure and urine 6-SMT (6-sulfatoxymelatonin, metabolism product of MLT in urine) in the daytime and nighttime were measured in normal control subjects (n = 20), non-dipping HPT group (n = 32) and dipping HPT group (n = 36). RESULTS: As expected, blood pressure reduction during the night was significantly lower in non-dipping HPT group compared to control and dipping HPT groups. Nighttime 6-SMT was significantly higher than daytime 6-SMT in all groups and nighttime urine 6-SMT levels and ratio of nighttime/daytime urine 6-SMT levels were significantly lower in non-dipping HPT group compared to control and dipping HPT groups (all P < 0.01). Moreover, ratio of nighttime/daytime urine 6-SMT levels are positively correlated blood pressure reduction during nighttime (P < 0.05) in all groups. CONCLUSIONS: Circadian rhythm of MLT secretion is maintained but nighttime MLT secretion was significantly reduced and related to disorders of circadian rhythm of blood pressure in non-dipping HPT group.

Damage to the nigrostriatal system in the MPTP-treated SAMP8 mouse.

Application of aged animals to studies of Parkinson's disease (PD) will be beneficial to improve the understanding of its pathogenesis. The senescence-accelerated mouse prone8 (SAMP8) mouse has an early onset of senility and a short life span, characterized by learning and memory impairment, and affective disturbance in the aging process. There is no animal currently being used as a PD model that exhibits these characteristics. Application of the SAMP8 mouse to PD research may have several merits. For the first time, we have investigated damage of the nigrostriatal system in the SAMP8 mouse induced by 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP). Male SAMP8 mice (12 weeks) were treated with four subcutaneous injections of MPTP (20mg/kg at 2h intervals): spontaneous activity decreased significantly after the third injection, and recovered 48h after the first injection. In MPTP-SAMP8 mice, the tyrosine hydroxylase (TH)-positive neuronal loss at 6h (7.06%), 24h (12.79%), 3 days (22.49%), and 8 days (42.39%), while striatal dopamine (DA) levels decreased at 6h by 79.09%, at 24h by 80.33%, at 3 days by 83.86%, and at 8 days by 80.14%. These results indicated that there were marked decreases in striatal DA levels and a loss of dopaminergic neurons in the substantia nigra, with the behavior change following shortly thereafter, in MPTP-SAMP8 mice. On the basis of the current findings, the SAMP8 mouse is also vulnerable to neurotoxic effects of MPTP. These data suggest that the SAMP8 mouse may be utilized in PD research.