Glia-to-Neuron Conversion by CRISPR-CasRx Alleviates Symptoms of Neurological Disease in Mice.

Conversion of glial cells into functional neurons represents a potential therapeutic approach for replenishing neuronal loss associated with neurodegenerative diseases and brain injury. Previous attempts in this area using expression of transcription factors were hindered by the low conversion efficiency and failure of generating desired neuronal types in vivo. Here, we report that downregulation of a single RNA-binding protein, polypyrimidine tract-binding protein 1 (Ptbp1), using in vivo viral delivery of a recently developed RNA-targeting CRISPR system CasRx, resulted in the conversion of Müller glia into retinal ganglion cells (RGCs) with a high efficiency, leading to the alleviation of disease symptoms associated with RGC loss. Furthermore, this approach also induced neurons with dopaminergic features in the striatum and alleviated motor defects in a Parkinson's disease mouse model. Thus, glia-to-neuron conversion by CasRx-mediated Ptbp1 knockdown represents a promising in vivo genetic approach for treating a variety of disorders due to neuronal loss.

[Effect of bilateral superior cervical sympathetic ganglion occlusion on pathological process of aortic dissection and its mechanism].

OBJECTIVE: To investigate the effect of bilateral superior cervical sympathetic ganglion occlusion (SCG) on aortic dissection and its possible mechanism. METHODS: Forty-five SD rats were randomly divided into three groups with 15 in each:blank control group, sham operation group and SCG group. ß-aminopropione (666 mg·kg-1·d-1) was given by subcutaneous injection for 4 weeks to establish the aortic dissection model. Rats in SCG group were given SCG before the injection of ß-aminopropione. Blood pressure and heart rate of the rats were monitored using noninvasive tail artery blood pressure measuring instrument; sympathetic activity was monitored using drug block method; the structure of aortic wall was observed using HE staining; collagen fibers in aortic wall was observed using Sirius red staining; protein expression of Apelin was detected by immunohistochemistry; and the protein expression of matrix metalloproteinase (MMP)-2, 9 was detected by Western blotting. RESULTS: During the experiment, the body mass of the sham operation group and SCG group was smaller than that of the blank control group (all P<0.05), and the body mass of the SCG group was larger than that of the sham operation group (all P<0.05). The heart rate and sympathetic activity of the sham operation group were higher than those of the blank control group (all P<0.05), while the SCG group were lower (all P<0.05). Compared with the blank control group, the aortic wall in the sham operation group was thickening, while that in the SCG group was improved. A large number of collagen-1 in the aortic wall of the blank control group was stained brown by Sirius red, which was lighter in SCG group, and the staining in the sham operation group was the lightest. Compared with the blank control group, the expression of Apelin, MMP-2 and MMP-9 protein in the sham operation group increased (all P<0.05), while those in the SCG group decreased (all P<0.05). CONCLUSIONS: SCG can effectively reduce the incidence and mortality of aortic dissection in rats, which may be related to the inhibition of sympathetic activity and the decrease of collagen-1, Apelin, MMP-2 and MMP-9 expression.

[The number and subgroups of lymphocytes in valve tissue of rheumatic heart disease combined with diabetes mellitus].

Objective To investigate the effect of diabetes mellitus on lymphocytes in rheumatic heart valve tissue and its mechanism. Methods Valve tissues of 40 patients undergoing heart valve replacement were collected, including 20 patients in rheumatic heart disease group (without diabetes) and 20 patients in diabetic group (rheumatic heart disease combined with diabetes). In addition, 20 cases of valve tissue from control group were collected. HE staining was used to observe the damage of valve tissue and the area of collagen degeneration. CD4+ T cells, CD8+ T cells, B cells and plasma cells were detected by immunohistochemical staining. Flow cytometry was used to detect the proportion of regulatory T cells (Tregs) in peripheral blood. Results Compared with the rheumatic heart disease group, the damage of valve tissue in the diabetic group was further aggravated, the number of infiltrating inflammatory cells increased, and the area of collagen degeneration was enlarged. Compared with the control group, the number of T cells, CD4+ T cells, CD8+ T cells, B cells and plasma cells in valve tissue of patients with rheumatic heart disease increased significantly. Diabetes mellitus further increased the number of T cells, CD4+ T cells, B cells and plasma cells in valve tissue, but had no significant effect on CD8+ T cells. The proportion of Tregs in the peripheral blood of patients with rheumatic heart disease was significantly reduced. Diabetes mellitus could further reduce the proportion of Tregs. Conclusion The number of T cells, CD4+T cells, B cells and plasma cells in heart valves of rheumatic heart disease patients with diabetes mellitus go up significantly, and Treg ratio goes down.

Reduction of HIP2 expression causes motor function impairment and increased vulnerability to dopaminergic degeneration in Parkinson's disease models.

Huntingtin interaction protein 2 (HIP2) is an E2 ubiquitin-conjugating enzyme associated with neurodegenerative diseases, and HIP2 mRNA has been implicated as a potential blood biomarker for Parkinson's disease (PD). However, it is unclear whether the alteration of HIP2 expression may contribute to the development of PD, and whether the change of HIP2 in blood could reflect its expression in the brain or motor functions in PD patients. In this study, we established a mouse line with HIP2 haploinsufficiency. The reduction of the HIP2 expression led to spontaneous motor function impairment and dopaminergic neuronal loss. Furthermore, HIP2 haploinsufficiency increased the susceptibility of mice to 6-hydroxydopamine (6-OHDA) and caused severe loss of dopaminergic neurons. Interestingly, in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model for PD, we observed concurrent, highly correlated decrease of HIP2 expression in the brain and in the blood. Using blood samples from more than 300 patients, we validated the decreased HIP2 mRNA in PD patients, including de novo patients. Finally, in a 1-year, 20-patient study, we observed reversed blood HIP2 mRNA levels accompanying improved motor and overall daily functions in 75% of the PD patients with instructed Tai Chi training. Therefore, our in vivo studies have indicated HIP2 insufficiency as a contributing factor for PD, and functionally validated blood HIP2 as a useful and reversible biomarker for PD.

Chymase induces profibrotic response via transforming growth factor-beta 1/Smad activation in rat cardiac fibroblasts.

Mast cell-derived chymase is implicated in myocardial fibrosis (MF), but the underlying mechanism of intracellular signaling remains unclear. Transforming growth factor-beta 1 (TGF-beta1) is identified as the most important profibrotic cytokine, and Smad proteins are essential, but not exclusive downstream components of TGF-beta 1 signaling. Moreover, novel evidence indicates that there is a cross talk between Smad and mitogen-activated protein kinase (MAPK) signaling cascade. We investigated whether chymase activated TGF-beta 1/Smad pathway and its potential role in MF by evaluating cardiac fibroblasts (CFs) proliferation and collagen synthesis in neonatal rats. MTT assay and 3H-Proline incorporation revealed that chymase induced CFs proliferation and collagen synthesis in a dose-dependent manner. RT-PCR and Western blot assay demonstrated that chymase not only increased TGF-beta1 expression but also upregulated phosphorylated-Smad2/3 protein. Furthermore, pretreatment with TGF-beta 1 neutralizing antibody suppressed chymase-induced cell growth, collagen production, and Smad activation. In contrast, the blockade of angiotensin II receptor had no effects on chymase-induced production of TGF-beta 1 and profibrotic action. Additionally, the inhibition of MAPK signaling had no effect on Smad activation elicited by chymase. These results suggest that chymase can promote CFs proliferation and collagen synthesis via TGF-beta 1/Smad pathway rather than angiotensin II, which is implicated in the process of MF.