Novel cryptic exons identified in introns 2 and 3 of the human dystrophin gene with duplication of exons 8-11

The dystrophin gene, which is mutated in Duchenne muscular dystrophy, is thelargest known human gene and characterized by the huge size of its introns. Intron 2has been shown to include cryptic exons termed exons 2a and 2b, while intron 3 hasbeen shown to include a cryptic exon designated exon 3a. In the present study, weidentified 2 and 1 additional cryptic exons in introns 2 and 3, respectively. A previouslyunknown 157-bp insertion was identified between exons 2 and 3 of a dystrophin mRNAisolated from the lymphocytes of a dystrophinopathy patient with duplication of exons8-11. Since this sequence exhibited the typical characteristics of a genomic exon, wedesignated it 'exon 2c-l'. A more detailed examination revealed that a position 4 bpdownstream from the 5' end of exon 2c-l was also used as a splice acceptor site, and thisexon was designated 'exon 2c-s'. In the same patient, a 357-bp insertion was identifiedbetween exons 3 and 4. Since this sequence also showed the typical characteristics of anexon, and its 3' end was the same as the splice donor site of exon 3a, we designated thenovel cryptic exon 'exon 3a-l', and changed the name of the previously reported exon3a to 'exon 3a-s'. Among these novel cryptic exons, exon 3a-l was also incorporatedinto the dystrophin mRNA from normal lymphocytes, whereas exons 2c-l and 2c-s werenot. The physiological or pathophysiological roles of these novel cryptic exons remainto be clarified

頸椎症性脊髄症における椎間関節変性の有病率と臨床的影響：CT新分類での検討

Objective: To evaluate cervical facet joint degeneration using a newly developed classification, investigate its prevalence and relationship with cervical degenerative spondylolisthesis, and clarify its clinical significance in patients with degenerative cervical myelopathy (DCM). Methods: This study included 145 consecutive patients with DCM who underwent surgical treatment. Clinical variables and radiological findings were analyzed. A new 6-grade computed tomography (CT) classification for cervical facet joint degeneration was adapted, and its prevalence was evaluated by categorizing the joints into those at responsible and those at nonresponsible spinal segmental levels. We evaluated the association between rapidly progressive myelopathy and the presence of significant facet joint degeneration or spondylolisthesis at the responsible segmental level. Results: Finally, 140 patients with a mean age of 64.1±12.8 years were analyzed. The prevalence of grade 1, 2, 3, 4, 5A, and 5B classification in all facet joints was 72.0%, 9.5%, 10.9%, 4.3%, 2.9%, and 0.4%, respectively. There was a statistically significant difference in the distribution of CT grades between the joints at the responsible and nonresponsible segmental levels (p<0.001), with a high prevalence of grade 4 or 5B degeneration at the responsible segmental level, reflecting articular irregularity. There was also a statistically significant relationship between rapidly progressive myelopathy and grade 4 or 5B degeneration at the responsible segmental level (p<0.001), but not between rapidly progressive myelopathy and spondylolisthesis (p=0.255). Conclusion: This novel CT classification for facet joints deserves additional evaluation in patients with DCM. Abnormal findings on the articular surfaces might be related to the progression of myelopathy.博士（医学）・甲第870号・令和5年3月15

Cilostazol minimizes venous ischemic injury in diabetic and normal rats

We evaluated the effects of cilostazol on venous infarction produced by a photothrombotic two-vein occlusion (2VO) model in diabetic and control rats. The cerebral blood flow (CBF) between the occluded veins was measured by laser Doppler flowmetry for 4 hours after 2VO. Infarct size and immunohistochemistry were evaluated 24, 48, 96, and 168 hours after 2VO. Cilostazol was administered 1 hour after 2VO, and thereafter at a continuous oral dose of 60 mg/kg per day. Cilostazol reduced the infarct size, and the number of terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL)-positive apoptotic and B-cell lymphoma 2-associated X protein (Bax)-positive cells, and improved the CBF in control rats. In diabetic rats, cilostazol reduced the infarct size, and the number of TUNEL-positive apoptotic and Bax-positive cells, 96 and 168 hours after 2VO, but did not improve the CBF 4 hours after 2VO. Cilostazol increased the number of B-cell lymphoma 2 (Bcl-2)-positive cells in both strains 48, 96, and 168 hours after 2VO, but did not improve vessel wall thickness or collagen deposits. Cilostazol appeared to limit venous infarcts by improving the penumbral CBF in nondiabetic rats, and inhibited pro-apoptotic changes through Bcl-2 overexpression, without improving the CBF in diabetic rats

脳皮質静脈梗塞ラットモデルを用いた脳静脈虚血におけるプラバスタチンの神経保護効果

Objectives: Pravastatin sodium is reported to have multiple beneficial effects in cerebral atherosclerosis and neuronal injury; however, the preventive effects on cerebral venous ischemia are still unknown. Herein, we aimed to examine the neuroprotective effects of transoral prior administration of pravastatin sodium against cerebral cortical venous ischemia with suppression of apoptosis. Methods: Thirty 8-week-old male Wistar rats were divided equally into two study groups (n = 15 vs. n = 15); the pravastatin group was fed 1% pravastatin sodium with their usual diet for 2 weeks, while the control group only received the usual diet. Two-vein occlusion (2VO) model was applied for this study, and two adjacent cortical veins in each animal were permanently occluded photochemically with rose bengal dye. During photothrombosis, regional changes of the cerebral blood flow (CBF) in area of the venous ischemia were recorded. At 48-h after 2VO, animals were euthanized using perfusion fixation, and we histologically measured ratios of infarcted area to contralateral hemisphere, and counted Bax- and Bcl-2-positive cells in the penumbra to investigate the implications for apoptosis. Results: The ratio of infarcted area was significantly decreased in the pravastatin group compared to the control group (P < 0.01). The number of Bax-positive cells also decreased significantly in the pravastatin group (P < 0.01). In contrast, immunolabeling for Bcl-2 was essentially negative in all areas in both groups. There were also no significant differences in regional CBF changes after 2VO between the two groups (P = 0.13). Conclusions: Pre-emptive administration of pravastatin sodium mixed in the food has neuroprotective effects against cerebral cortical venous ischemia with suppression of apoptosis associated with inhibition of Bax expression but has little influence on regional CBF.権利情報：© 2023 The Authors. Published by Elsevier Ltd on behalf of International Brain Research　Organization. This is an open access article under the CC BY　license　(http://creativecommons.org/licenses/by/4.0/)

脳皮質静脈梗塞ラットモデルを用いた脳静脈虚血による神経細胞および神経前駆細胞の発現と分布

Neurogenesis in the subventricular zone (SVZ), subgranular zone (SGZ), and cerebral cortex is now a familiar event to confirm by cerebral arterial ischemia in rat models. However, it remains unclear whether cerebral venous ischemia (CVI) alone causes neurogenesis, and where that neurogenesis occurs. After creating CVI rat models via a two-vein occlusion (2-VO) method, neurogenesis was immunohistochemically evaluated by doublelabeling 5-bromo-2′ -deoxyuridine (BrdU)-positive cells with neuronal nuclei (NeuN) or doublecortin (DCX) antibody. Fifty Wistar rats were divided into two major groups (BrdU-NeuN and BrdU-DCX) and then separated into two subgroups (2-VO or sham). The total number of double-positive cells expressed inside a predefined region of interest (ROI) covering the ischemic area was compared between the two subgroups. Then, we divided the ROI into six sections to evaluate and compare the distribution of double-positive cells generated in each section between the two subgroups. The 2-VO subgroup presented more double-positive cells than the sham group in both BrdU-NeuN and BrdU-DCX groups, while the BrdU-DCX+2-VO group showed a characteristic distribution of double-positive cells in ROI 2 and ROI 3, suggesting areas of the ischemic core and penumbra, with a significant difference compared to the BrdU-DCX+sham group. This study demonstrates that CVI has the potential to induce endogenous neurogenesis, with significant numbers of both newly generated neurons and precursors observed in the ischemic area. The distribution of these cells suggests that the cortex could be the main origin of neurogenesis after cortical CVI.博士（医学）・甲第872号・令和5年3月15

On the Nature of AX J2049.6+2939 and AX J2050.0+2914

AX J2049.6+2939 is a compact X-ray source in the vicinity of the southern blow-up region of the Cygnus Loop supernova remnant (Miyata et al. 1998a). This source was the brightest X-ray source inside the Cygnus Loop observed during the ASCA survey project. The X-ray spectrum was well fitted by a power-law function with a photon index of $-2.1 \pm 0.1$. Short-term timing analysis was performed and no coherent pulsation was found. Follow-up observations with ASCA have revealed a large variation in X-ray intensity by a factor of $\simeq$ 50, whereas the spectral shape did not change within the statistical uncertainties. In the second ASCA observation, we found another X-ray source, AX J2050.0+2941, at the north east of AX J2049.6+2939. During the three ASCA observations, the X-ray intensity of AX J2050.0+2941 varied by a factor of $\simeq$4. No coherent pulsations could be found for AX J2050.0+2941. We have performed optical photometric and spectroscopic observations in the vicinity of AX J2049.6+2939 at the Kitt Peak National Observatory (KPNO). As a result, all objects brighter than $B$-band magnitude of 22 in the error box can be identified with normal stars. Combined with the X-ray results and the fact that there are no radio counterparts, AX J2049.6+2939 is not likely to be either an ordinary rotation-powered pulsar or an AGN. The nature of AX J2049.6+2939 is still unclear and further observations over a wide energy band are strongly required. As to AX J2050.0+2941, the long-term X-ray variability and the radio counterpart suggests that it is an AGN.Comment: 23 pages, 4 figures, Accepted for publication by Astrophysical Journa