Salt-induced hypertension in WKY rats: Prevention by α-lipoic acid supplementation

There is strong evidence that points to excess dietary salt as a major factor contributing to the development of hypertension. Salt sensitivity is associated with glucose intolerance and insulin resistance in both animal models and humans. In insulin resistance, impaired glucose metabolism leads to elevated endogenous aldehydes which bind to vascular calcium channels, increasing cytosolic [Ca²⁺]_i and blood pressure. In an insulin resistant animal model of hypertension, spontaneously hypertensive rats (SHRs), dietary supplementation with lipoic acid lowers tissue aldehydes and plasma insulin levels and normalizes blood pressure. The objective of this study is to examine the effects of a high salt diet on tissue aldehydes, cytosolic [Ca²⁺]_i and blood pressure in WKY rats and to investigate whether dietary supplementation with lipoic acid can prevent a salt induced increase in blood pressure. Starting at 7 weeks of age, WKY rats were divided into three groups of six animals each and treated for 10 weeks with diets as follows: WKY-normal salt (0.7% NaCl); WKY-high salt (8% NaCl); WKY-high salt + lipoic acid (8% NaCl diet + lipoic acid 500 mg/Kg feed). At completion, animals in the high salt group had elevated systolic blood pressure, platelet [Ca²⁺]_i, and tissue aldehyde conjugates compared with the normal salt group and showed smooth muscle cell hyperplasia in the small arteries and arterioles of the kidneys. Dietary -lipoic acid supplementation in high salt-treated WKY rats normalized systolic blood pressure and cytosolic [Ca²⁺]_i and aldehydes in liver and aorta. Kidney aldehydes and renal vascular changes were attenuated, but not normalized.     

The Wilms Tumor Gene,Wt1, Is Critical for Mouse Spermatogenesis via Regulation of Sertoli Cell Polarity and Is Associated with Non-Obstructive Azoospermia in Humans

Azoospermia is one of the major reproductive disorders which cause male infertility in humans; however, the etiology of this disease is largely unknown. In the present study, six missense mutations of WT1 gene were detected in 529 human patients with non-obstructive azoospermia (NOA), indicating a strong association between WT1 mutation and NOA. The Wilms tumor gene, Wt1, is specifically expressed in Sertoli cells (SCs) which support spermatogenesis. To examine the functions of this gene in spermatogenesis, Wt1 was deleted in adult testis using Wt1^flox and Cre-ER^TM mice strains. We found that inactivation of Wt1 resulted in massive germ cell death and only SCs were present in most of the seminiferous tubules which was very similar to NOA in humans. In investigating the potential mechanism for this, histological studies revealed that the blood–testis barrier (BTB) was disrupted in Wt1 deficient testes. In vitro studies demonstrated that Wt1 was essential for cell polarity maintenance in SCs. Further studies found that the expression of cell polarity associated genes (Par6b and E-cadherin) and Wnt signaling genes (Wnt4, Wnt11) were downregulated in Wt1 deficient SCs, and that the expression of Par6b and E-cadherin was regulated by Wnt4. Our findings suggest that Wt1 is important in spermatogenesis by regulating the polarity of SCs via Wnt signaling pathway and that WT1 mutation is one of the genetic causes of NOA in humans.     

Early Postnatal EEG Features of Perinatal Arterial Ischaemic Stroke with Seizures

Background

Stroke is the second most common cause of seizures in term neonates and is associated with abnormal long-term neurodevelopmental outcome in some cases.

Objective

To aid diagnosis earlier in the postnatal period, our aim was to describe the characteristic EEG patterns in term neonates with perinatal arterial ischaemic stroke (PAIS) seizures.

Design

Retrospective observational study.

Patients

Neonates >37 weeks born between 2003 and 2011 in two hospitals.

Method

Continuous multichannel video-EEG was used to analyze the background patterns and characteristics of seizures. Each EEG was assessed for continuity, symmetry, characteristic features and sleep cycling; morphology of electrographic seizures was also examined. Each seizure was categorized as electrographic-only or electroclinical; the percentage of seizure events for each seizure type was also summarized.

Results

Nine neonates with PAIS seizures and EEG monitoring were identified. While EEG continuity was present in all cases, the background pattern showed suppression over the infarcted side; this was quite marked (>50% amplitude reduction) when the lesion was large. Characteristic unilateral bursts of theta activity with sharp or spike waves intermixed were seen in all cases. Sleep cycling was generally present but was more disturbed over the infarcted side. Seizures demonstrated a characteristic pattern; focal sharp waves/spike-polyspikes were seen at frequency of 1–2 Hz and phase reversal over the central region was common. Electrographic-only seizure events were more frequent compared to electroclinical seizure events (78 vs 22%).

Conclusions

Focal electrographic and electroclinical seizures with ipsilateral suppression of the background activity and focal sharp waves are strong indicators of PAIS. Approximately 80% of seizure events were the result of clinically unsuspected seizures in neonates with PAIS. Prolonged and continuous multichannel video-EEG monitoring is advocated for adequate seizure surveillance.     

Somaclonal variation does not preclude the use of rice transformants for genetic screening

Rice (Oryza sativa) is one of the world's most important crops. Rice researchers make extensive use of insertional mutants for the study of gene function. Approximately half a million flanking sequence tags from rice insertional mutant libraries are publicly available. However, the relationship between genotype and phenotype is very weak. Transgenic plant assays have been used frequently for complementation, overexpression or antisense analysis, but sequence changes caused by callus growth, Agrobacterium incubation medium, virulence genes, transformation and selection conditions are unknown. We used high‐throughput sequencing of DNA from rice lines derived from Tainung 67 to analyze non‐transformed and transgenic rice plants for mutations caused by these parameters. For comparison, we also analyzed sequence changes for two additional rice varieties and four T‐DNA tagged transformants from the Taiwan Rice Insertional Mutant resource. We identified single‐nucleotide polymorphisms, small indels, large deletions, chromosome doubling and chromosome translocations in these lines. Using standard rice regeneration/transformation procedures, the mutation rates of regenerants and transformants were relatively low, with no significant differences among eight tested treatments in the Tainung 67 background and in the cultivars Taikeng 9 and IR64. Thus, we could not conclusively detect sequence changes resulting from Agrobacterium‐mediated transformation in addition to those caused by tissue culture‐induced somaclonal variation. However, the mutation frequencies within the two publically available tagged mutant populations, including TRIM transformants or Tos17 lines, were about 10‐fold higher than the frequency of standard transformants, probably because mass production of embryogenic calli and longer callus growth periods were required to generate these large libraries.     

Water-soluble quantum dots for biomedical applications

Semiconductor nanocrystals are 1-10nm inorganic particles with unique size-dependent optical and electrical properties due to quantum confinement (so they are also called quantum dots). Quantum dots are new types of fluorescent materials for biological labeling with high quantum efficiency, long-term photostability, narrow emission, and continuous absorption spectra. Here, we discuss the recent development in making water-soluble quantum dots and related cytotoxicity for biomedical applications.     

Retinoic acid–induced survival effects in SH-SY5Y neuroblastoma cells

Neuroblastoma is a malignant childhood cancer arising from the embryonic sympathoadrenal lineage of the neural crest. Retinoic acid (RA) is included in the multimodal therapy of patients with high-risk neuroblastoma to eliminate minimal residual disease. However, the formation of RA-resistant cells substantially lowers 5-year overall survival rates. To examine mechanisms that lead to treatment failure, we chose human SH-SY5Y cells, which are known to tolerate incubation with RA by activating the survival kinases Akt and extracellular signal-regulated kinase 1/2. Characterization of downstream pathways showed that both kinases increased the phosphorylation of the ubiquitin ligase mouse double minute homolog 2 (Mdm2) and thereby enhanced p53 degradation. When p53 signaling was sustained by blocking complex formation with Mdm2 or enhancing c-Jun N-terminal kinase (JNK) activation, cell viability was significantly reduced. In addition, Akt-mediated phosphorylation of the cell-cycle regulator p21 stimulated complex formation with caspase-3, which also contributed to cell protection. Thus, treatment with RA augmented survival signaling and attenuated basal apoptotic pathways in SH-SY5Y cells, which increased cell viability.     

Corneal Changes and Strategies to Improve Survival of Hypomorphic Collagen VII-Deficient Mice for the Study of Ocular Dystrophic Epidermolysis Bullosa

Ophthalmic study of collagen CVII hypomorphic mice is uniquely challenging due to the strain’s published survival rate to weaning of 24%. Because chronic ocular fibrosis requires time to develop, optimizing the survival rate is of critical importance. In this study, standard husbandry practices were enhanced by the addition of sterilized diet and drug delivery gels, acidified water, irradiated food pellets, cellulose fiber bedding, minimal handling, removal of siblings within 2-3 wk from birth, and a preferred housing location. Survival rates per breeding cycle, sex, weight, and cause of early euthanasia were recorded and analyzed over 43 mo. Overall, 49% of mice survived to weaning and 76% of weaned mice survived to 20 wk of age. Corneal opacities were seen in 65% of mice by 20 wk, but only 10% of eyes showed the sustained opacification that was indicative of fibrosis. Corneal opacities occurred at the same rate as in humans with epidermolysis bullosa. 66% of the mice showed weight loss at 11 wk. Males required early euthanasia 4 times more often than did females. Euthanasia was required for urinary obstruction due to penile prolapse in 88% of males. With our enhanced care protocol, hypomorphic mice in our colony survived at twice the published rate. With this revised husbandry standard, experiments planned with termination endpoints of 14 wk for males and 17 wk for females are more likely to reach completion.