The mitochondrial toxin 3-nitropropionic acid induces differential expression patterns of apoptosis-related markers in rat striatum

The mitochondrial toxin 3-nitropropionic acid (3-NP) causes selective striatal lesions in rats and serves as an experimental model for the neurodegenerative disorder Huntington's disease (HD). Apoptotic cell death has been implicated for the neuronal degeneration that occurs in HD brains. The present study was designed to investigate whether the 3-NP-induced cell death in rats involves apoptosis and an altered expression of Bcl-2 family proteins. Systemic administration of 3-NP via subcutaneous Alzet pumps resulted in lesions of variable severity with neuronal loss and gliosis in the striatum. Using the terminal transferase-mediated biotinylated-UTP nick end-labelling (TUNEL) of DNA, TUNEL-positive cells exhibiting typical apoptotic morphology were detected only in the striatum of rats with a severe lesion. Furthermore, the neuronal expression of the pro-apoptotic protein Bax was strongly increased in the core of the severe lesion. Expression of the anti-apoptotic marker Bcl-2 was unchanged in this location, but was enhanced in the margins of the lesions. A moderately increased expression of both Bax and Bcl-2 was observed in dark neurones in the mild lesion and in the subtle lesion. The presence of nuclear DNA fragmentation, strong granular Bax expression and an increased Bax/Bcl-2 ratio in the centre of severe lesions suggests the occurrence of apoptotic cell death following 3-NP administration. In contrast, the dark compromised neurones observed in 3-NP-treated animals revealed an equally enhanced expression of both Bax and Bcl-2, but lacked TUNEL-labelling, and are therefore not apoptotic.     

44Sacral extracorporeal magnetic stimulation is an effective treatment for pelvic floor dyssynergia; Comparative study with biofeedback therapy

Background: Recent development of extracorporeal magnetic stimulation (ECMS) which uses current‐changing magnetic fields allows the induction of electrical stimulation in the desired deep tissue. Recent study showed the sacral nerve stimulation reduces corticoanal excitability that may play a functional role in anal continence mechanisms. Preliminary study shows that ECMS of sacral nerve can modify pelvic floor function and expel rectal balloon in patients with pelvic floor dyssynergia (PFD). Aims: To evaluate the effect of ECMS compared with biofeedback therapy (BF) in patients with PFD. Methods and Materials: Thirty‐eight patients who fulfilled Rome II criteria for PFD by colon transit time and anorectal function tests, were randomly treated with 8 sessions of ECMS (2/weeks; n = 19) at prone position or BF (2/weeks; n = 19) at sitting position. Stimulation parameters were set at 50–80% of maximum intensity, 10 and 50 Hz frequency, 3 s burst length with 3 and 6 s off using arm‐typed stimulator (BioCom‐1000, Mcube Co., Korea). Symptom scores for constipation with/without anorectal function test were repeatedly measured after each treatment. Response was defined as 50% or more decreased symptom score after treatment (partial response: 30–50%, poor: <30%). Results: Fifteen patients (age 49.1 ± 13.4 years, mean ± SD; 4 men) completed 8 session of BF and 14 patients (54.5 ± 17.6 years, 3 men) completed 8 session of ECMS. Four patients of BF group discontinued treatment due to unsatisfactory therapeutic effect (n = 1) and withdrew consent (n = 3) and 5 patients of ECMS group discontinued treatment because of same reasons (n = 1, 4). Total symptom scores were significantly decreased after treatment of 8 session in both treatment groups (13.4 ± 6.6 vs. 4.3 ± 4.0 for BF, p = 0.009; 14.9 ± 5.6 vs. 3.4 ± 4.0 for ECMS, p < 0.001). Bowel movements per week were also significantly increased after treatment in both groups (median 2 vs. 7 for BF, p = 0.035; median 2 vs. 7 for ECMS, p = 0.008). Thirteen out of 15 patients showed response in BF group and 12 out of 14 showed good response in ECMS group. No adverse effects in both groups. Conclusions: ECMS is as effective as BF for the treatment of PFD. Long‐term effect of ECMS for the patients with pelvic floor dyssynergia need to be evaluated in the near future.     

Functional analysis of novel KCNQ2 mutations found in patients with Benign Familial Neonatal Convulsions

Benign Familial Neonatal Convulsions (BFNC) are a rare epilepsy disorder with an autosomal-dominant inheritance. It is linked to mutations in the potassium channel genes KCNQ2 and KCNQ3. These encode for Kv7.2 and Kv7.3 potassium ion channels, which produce an M-current that regulates the potential firing action in neurons through modulation of the membrane potential. We report on the biophysical and biochemical properties of V589X, T359K and P410fs12X mutant-KCNQ2 ion channels that were detected in three BFNC families. Mutant KCNQ2 cDNAs were co-expressed with WT-KCNQ2 and KCNQ3 cDNAs in HEK293 cells to mimic heterozygous expression of the KCNQ2 mutations in BFNC patients. The resulting potassium currents were measured using patch-clamp techniques and showed an approximately 75% reduction in current and a depolarized shift in the voltage dependence of activation. Furthermore, the time-constant of activation of M-currents in cells expressing T359K and P410fs12X was slower compared to cells expressing only wild-type proteins. Immunofluorescent labeling of HEK293 cells stably expressing GFP-tagged KCNQ2-WT or mutant α-subunits indicated cell surface expression of WT, V589X and T359K mutants, suggesting a loss-of-function, while P410fs12X was predominantly retained in the ER and sub-cellular compartments outside the ER suggesting an effectively haplo-insufficient effect.     

AMP-18,一种新发现的胃黏膜保护因子

AMP-18是一种新发现的由胃腺体上皮细胞合成的小分子蛋白质,独特表达于胃黏膜,机体其他部位少见,胃癌组织中表达缺失．AMP-18 由185个氨基酸组成,除去N端信号肽(20个氨基酸)后大小约18 ku,第54-150个氨基酸组成高度保守的结构域(BRICHOS区域)承担主要的生理功能．AMP-18由胃腺体上皮细胞以胞吐的方式分泌到胃黏液中,他的合成和分泌与个体生长发育有关,并受福斯高林、吲哚美辛、地塞米松等药物的影响．目前发现 AMP-18的生理功能主要有促进胃黏膜上皮细胞的有丝分裂,促进细胞的迁徙,促胃肠黏膜损伤的修复,保持胃肠黏膜的完整等．     

A founder mutation in BBS2 is responsible for Bardet‐Biedl syndrome in the Hutterite population: utility of SNP arrays in genetically heterogeneous disorders

Innes AM, Boycott KM, Puffenberger EG, Redl D, MacDonald IM, Chudley AE, Beaulieu C, Perrier R, Gillan T, Wade A, Parboosingh JS. A founder mutation in BBS2 is responsible for Bardet‐Biedl syndrome in the Hutterite population: utility of SNP arrays in genetically heterogeneous disorders. Bardet‐Biedl syndrome (BBS) is a multisystem genetically heterogeneous disorder, the clinical features of which are largely the consequence of ciliary dysfunction. BBS is typically inherited in an autosomal recessive fashion, and mutations in at least 14 genes have been identified. Here, we report the identification of a founder mutation in the BBS2 gene as the cause for the increased incidence of this developmental disorder in the Hutterite population. To ascertain the Hutterite BBS locus, we performed a genome‐wide single nucleotide polymorphism (SNP) analysis on a single patient and his three unaffected siblings from a Hutterite family. The analysis identified two large SNP blocks that were homozygous in the patient but not in his unaffected siblings, one of these regions contained the BBS2 gene. Sequence analysis and subsequent RNA studies identified and confirmed a novel splice site mutation, c.472‐2A>G, in BBS2. This mutation was also found in homozygous form in three subsequently studied Hutterite BBS patients from two different leuts, confirming that this is a founder mutation in the Hutterite population. Further studies are required to determine the frequency of this mutation and its role, if any, in the expression of other ciliopathies in this population.     

Sleep misperception,EEG characteristics and Autonomic Nervous System activity in primary insomnia: A retrospective study on polysomnographic data

Misperception of Sleep Onset Latency, often found in Primary Insomnia, has been cited to be influenced by hyperarousal, reflected in EEG- and ECG-related indices. The aim of this retrospective study was to examine the association between Central Nervous System (i.e. EEG) and Autonomic Nervous System activity in the Sleep Onset Period and the first NREM sleep cycle in Primary Insomnia (n = 17) and healthy controls (n = 11). Furthermore, the study examined the influence of elevated EEG and Autonomic Nervous System activity on Stage2 sleep-protective mechanisms (K-complexes and sleep spindles). Confirming previous findings, the Primary Insomnia-group overestimated Sleep Onset Latency and this overestimation was correlated with elevated EEG activity. A higher amount of beta EEG activity during the Sleep Onset Period was correlated with the appearance of K-complexes immediately followed by a sleep spindle in the Primary Insomnia-group. This can be interpreted as an extra attempt to protect sleep continuity or as a failure of the sleep-protective role of the K-complex by fast EEG frequencies following within one second. The strong association found between K-alpha (K-complex within one second followed by 8–12 Hz EEG activity) in Stage2 sleep and a lower parasympathetic Autonomic Nervous System dominance (less high frequency HR) in Slow-wave sleep, further assumes a state of hyperarousal continuing through sleep in Primary Insomnia.     

Small heat shock proteins associated with cerebral amyloid angiopathy of hereditary cerebral hemorrhage with amyloidosis (Dutch type) induce interleukin-6 secretion

In hereditary cerebral hemorrhage with amyloidosis of the Dutch type (HCHWA-D), severe cerebral amyloid angiopathy (CAA) is associated with an inflammatory reaction. Small heat shock proteins (sHsps) are molecular chaperones and association of HspB8 with CAA in HCHWA-D has been observed. The aims of this study were to investigate (1) if other sHsps are associated with the pathological lesions in HCHWA-D brains, (2) if the amyloid-beta protein (A beta) increases production of sHsps in cultured cerebral cells and (3) if sHsps are involved in the cerebral inflammatory processes in both Alzheimer's disease (AD) and HCHWA-D. We conclude that Hsp20, HspB8 and HspB2 are present in CAA in HCHWA-D, and that A beta did not affect cellular sHsps expression in cultured human brain pericytes and astrocytes. In addition, we demonstrated that Hsp20, HspB2 and HspB8 induced interleukin-6 production in cultured pericytes and astrocytes, which could be antagonized by dexamethasone, whereas other sHsps and A beta were inactive, suggesting that sHsps may be among the key mediators of the local inflammatory response associated with HCHWA-D and AD lesions.     

Muscle ultrasound density in human fetuses with spina bifida aperta

Background

In fetal spina bifida aperta (SBA), leg movements caudal to the meningomyelocele (MMC) are transiently present, but they disappear shortly after birth. Insight in the underlying mechanism could help to improve treatment strategies. In fetal SBA, the pathogenesis of neuromuscular damage prior to movement loss is still unknown. We reasoned that prenatal assessment of muscle ultrasound density (fetal-MUD) could help to reveal whether progressive neuromuscular damage is present in fetal SBA, or not.

Aim

To reveal whether prenatal neuromuscular damage is progressively present in SBA.

Patients/methods

In SBA fetuses (n = 6; 22-37 weeks gestational age), we assessed fetal-MUD in myotomes caudal to the MMC and compared measurements between myotomes cranial to the MMC and controls (n = 11; 17-36 weeks gestational age). Furthermore, we intra-individually compared MUD and muscle histology between the pre- and postnatal period.

Results

Despite persistently present fetal leg movements caudal to the MMC, fetal-MUD was higher caudal to the MMC than in controls (p < 0.05). Fetal-MUD caudal to the MMC did not increase with gestational age, whereas fetal-MUD in controls and cranial to the MMC increased with gestational age (p < 0.05). In 5 of 6 patients assessed, comparison between pre- and postnatal MUD and/or muscle histology indicated consistent findings.

Conclusions

In fetal SBA, persistent leg movements concur with stable, non-progressively increased fetal-MUD. These data may implicate that early postnatal loss of leg movements is associated with the impact of additional neuromuscular damage after the prenatal period.     

3-Nitropropionic acid induces cell death and mitochondrial dysfunction in rat corticostriatal slice cultures

Exposure of organotypic rat corticostriatal slice cultures to the mitochondrial toxin 3-nitropropionic acid (3-NP) resulted in concentration-dependent loss of cresylviolet-stained cells and increase of lactate dehydrogenase and lactate efflux into the culture medium, indicators for cell death and metabolic activity in the slices, respectively. The involvement of apoptosis in these slices was suggested by using the terminal transferase-mediated biotinylated-UTP nick end-labeling (TUNEL) technique, and immunohistochemistry for the apoptosis-related markers Bax and Bcl-2. In 3-NP-exposed slices, TUNEL-positive cells were observed in both the striatum and the cortex but in different forms: striatal neurons were either diffusely stained or showed nuclear fragmentation, cortical neurons only exhibiting nuclear fragmentation. In 3-NP-exposed slices, the pro-apoptotic protein Bax was abundantly expressed, whereas the anti-apoptotic protein Bcl-2 was not expressed in striatal neurons. We suggest that both apoptosis and necrosis are involved in the 3-NP-treated slices, apoptosis as well as necrosis in the striatum and apoptosis in the cortex.