An Optimization of AAV-82Q-Delivered Rat Model of Huntington’s Disease

Objective@#: No optimum genetic rat Huntington model both neuropathological using an adeno-associated virus (AAV-2) vector vector has been reported to date. We investigated whether direct infection of an AAV2 encoding a fragment of mutant huntingtin (AV2-82Q) into the rat striatum was useful for optimizing the Huntington rat model. @*Methods@#: We prepared ten unilateral models by injecting AAV2-82Q into the right striatum, as well as ten bilateral models. In each group, five rats were assigned to either the 2×1012 genome copies (GC)/mL of AAV2-82Q (×1, low dose) or 2×1013 GC/mL of AAV2-82Q (×10, high dose) injection model. Ten unilateral and ten bilateral models injected with AAV-empty were also prepared as control groups. We performed cylinder and stepping tests 2, 4, 6, and 8 weeks after injection, tested EM48 positive mutant huntingtin aggregates. @*Results@#: The high dose of unilateral and bilateral AAV2-82Q model showed a greater decrease in performance on the stepping and cylinder tests. We also observed more prominent EM48-positive mutant huntingtin aggregates in the medium spiny neurons of the high dose of AAV2-82Q injected group. @*Conclusion@#: Based on the results from the present study, high dose of AAV2-82Q is the optimum titer for establishing a Huntington rat model. Delivery of high dose of human AAV2-82Q resulted in the manifestation of Huntington behaviors and optimum expression of the huntingtin protein in vivo.

Rho Guanine Nucleotide Exchange Factor 4 (Arhgef4) Deficiency Enhances Spatial and Object Recognition Memory

Guanine nucleotide exchange factors (GEFs) play multiple functional roles in neurons. In a previous study, we reported that Arhgef4 (Rho guanine nucleotide exchange factor 4) functioned as a negative regulator of the excitatory synaptic function by sequestering postsynaptic density protein 95 (PSD-95). However, the role ofArhgef4 in behavior has not been examined. We performed comprehensive behavioral tests in knockout (KO) mice to investigate of the effects of Arhgef4 deficiency. We found that the expressed PSD-95 particle size was significantly increased in hippocampal neuronal cultures from Arhgef4 KO mice, which is consistent with the previous in vitro findings.Arhgef4 KO mice exhibited general motor activity and anxiety-like behavior comparable to those of the wild type littermates. However, spatial memory and object recognition memory were significantly enhanced in the Arhgef4 KO mice. Taken together, these data confirm the role of Arhgef4 as a negative synaptic regulator at the behavioral level.

Electrophysiological and Histologic Evaluation of the Time Course of Retinal Degeneration in the rd10 Mouse Model of Retinitis Pigmentosa

Among several animal models of retinitis pigmentosa (RP), the more recently developed rd10 mouse with later onset and slower rate of retinal degeneration than rd1 mouse is a more suitable model for testing therapeutic modalities. We therefore investigated the time course of retinal degeneration in rd10 mice before adopting this model in our interventional studies. Electroretinogram (ERG) recordings were carried out in postnatal weeks (PW) 3~5 rd10 (n=23) and wild-type (wt) mice (n=26). We compared the amplitude and implicit time of the b-wave of ERG records from wt and rd10 mice. Our results showed that b-wave amplitudes in rd10 mice were significantly lower and the implicit time of b-waves in rd10 mice were also significantly slower than that in wt mice (20~160 microV vs. 350~480 microV; 55~75 ms vs. 100~150 ms: p<0.001) through PW3 to PW5. The most drastic changes in ERG amplitudes and latencies were observed during PW3 to PW4. In multichannel recording of rd10 retina in PW2 to PW4.5, we found no significant difference in mean spike frequency, but the frequency of power spectral peak of local field potential at PW3 and PW3.5 is significantly different among other age groups (p<0.05). Histologic examination of rd10 retinae showed significant decrease in thickness of the outer nuclear layer at PW3. TUNEL positive cells were most frequently observed at PW3. From these data, we confirm that in the rd10 mouse, the most precipitous retinal degeneration occurs between PW3~PW4 and that photoreceptor degeneration is complete by PW5.

A Case of Idiopathic Erythrocytosis in a CAPD Patient / 대한신장학회잡지

We have experienced a case of idiopathic erythrocytosis developed in a patient with end stage renal disease who had switched to CAPD from hemodialysis. Hemoglobin levels gradually increased from 8 to 19.0 mg/dl, resulting in various symptoms from hyperviscosity during the first 2 months after the initiation of CAPD. There were no other possible causes of secondary erythrocytosis, such as hypoxia, erythropoietin -producing tumor or acquired cyst. Serum level of IGF-1 was above the normal range in contrast with low serum levels in CAPD patients with anemia. Increased IGF-1 levels may possibly influence on the development of erythrocytosis in this case.

beta PAK-interacting exchange factor may regulate actin cytoskeleton through interaction with actin

p21-activated kinase (PAK)-interacting exchange factor (PIX) is known to be involved in regulation of Cdc42/Rac GTPases and PAK activity. PIX binds to the proline-rich region of PAK, and regulates biological events through activation of Cdc42/Rac GTPase. To further investigate the role of PIX we produced monoclonal antibodies (Mab) against beta PAK. Three clones; N-C6 against N-terminal half and C-A3 and C-B7 against C- terminal half of beta PAK were generated and characterized. N-C6 Mab detected beta PAK as a major band in most cell lines. C-A3 Mab recognizes GIT-binding domain (GBD), but it does not interfere with GIT binding to beta PAK. Using C-A3 Mab possible beta PAK interaction with actin in PC12 cells was examined. beta PAK Mab (C-A3) specifically precipitated actin of the PC12 cell lysates whereas actin Mab failed to immunoprecpitate beta PAK. Co-sedimentation of PC12 cell lysates with the polymerized F-actin resulted in the recovery of most of beta PAK in the cell lysates. These results suggest that beta PAK may not interact with soluble actin but with polymerized F-actin and revealed that beta PAK constitutes a functional complex with actin. These data indicate real usefulness of the beta PAK Mab in the study of beta PAK role(s) in regulation of actin cyoskeleton.