Immunohistochemical investigation of nerve distribution in mature parotid and submandibular glands of rats with a liquid diet

Background: Although feeding with a liquid diet does not affect the growth of rat submandibular glands, it inhibits the growth of rat parotid glands during growth periods. In these growth-inhibited parotid glands, the growth of parasympathetic nerves is also suppressed. Meanwhile, the mature parotid glands of animals maintained on a liquid diet become morphologically and functionally atrophic, however, there is no effect of a liquid diet on mature submandibular glands. The objective of the present study was to clarify whether the nerve distribution in the mature salivary glands of rats was affected by a liquid diet. Materials and methods: Seven-week-old male Wistar rats were used in this study. Half of the rats were kept on a pellet diet, and half were kept on a liquid diet, for 3, 7, 14, or 21 days. All rats were euthanised by isoflurane at each endpoint. Then, the parotid and submandibular glands were removed, frozen in liquid nitrogen, cryosectioned, and stained with antibodies against protein gene product 9.5 (PGP 9.5; general nerve marker), tyrosine hydroxylase (TH; sympathetic nerve marker), or neuronal nitric oxide synthase (nNOS; parasympathetic nerve marker). Results: In parotid and submandibular glands of the pellet diet group, PGP 9.5- and TH-like immunoreactivity were seen around acini and ducts, and nNOS-like immunoreactivity was lower than PGP 9.5- and TH-like immunoreactivity. In the parotid glands of the liquid diet group, similar immunoreactivities were seen throughout the experimental period. The distribution of antibody labelling in the submandibular glands of the liquid diet group was similar to that of the pellet diet group and remained unchanged during the experimental period. Conclusions: The present study demonstrated no regressive effects of a liquid diet on the distribution of sympathetic or parasympathetic nerves in mature parotid glands and submandibular glands. This differed from inhibitory effects on the growth of parotid glands seen during growth periods

Update on the Keio collection of Escherichia coli single-gene deletion mutants

The Keio collection (Baba et al, 2006) has been established as a set of single‐gene deletion mutants of Escherichia coli K‐12. These mutants have a precisely designed deletion from the second codon from the seventh to the last codon of each predicted ORF. Further information is available at http://sal.cs.purdue.edu:8097/GB7/index.jsp or http://ecoli.naist.jp/. The distribution is now being handled by the National Institute of Genetics of Japan (http://www.shigen.nig.ac.jp/ecoli/pec/index.jsp). To date more than 4 million samples have been distributed worldwide. As we described earlier (Baba et al, 2006), gene amplification during construction is likely to have led to a small number of mutants with genetic duplications

High prevalence of wild-type transthyretin deposition in patients with idiopathic carpal tunnel syndrome: a common cause of carpal tunnel syndrome in the elderly

Carpal tunnel syndrome is the most common type of entrapment neuropathy. However, the cause of carpal tunnel syndrome remains unclear in most cases. Senile systemic amyloidosis, induced by wild-type transthyretin deposition, is a prevalent aging-related disorder and often accompanied by carpal tunnel syndrome. In this study, we measured the frequency of unrecognized wild-type transthyretin deposition in patients with idiopathic carpal tunnel syndrome. One hundred twenty-three patients with carnal tunnel syndrome, including 100 idiopathic patients, treated by carpal tunnel release surgery were analyzed. Tenosynovial tissues obtained at surgery were analyzed by Congo red and immunohistochemical staining. If staining for transthyretin was positive, the entire transthyretin gene was analyzed by direct DNA sequencing. We also analyzed tenosynovial tissues from 32 autopsy cases as controls. Thirty-four patients (34.0%) with idiopathic carpal tunnel syndrome showed amyloid deposition in the tenosynovial tissue, and all amyloid showed specific immunolabeling with antitransthyretin antibody. Direct DNA sequencing of the entire transthyretin gene did not reveal any mutations, indicating that all amyloid deposits were derived form wild-type transthyretin. Statistical analysis using logistic regression showed that the prevalence of transthyretin deposition in the idiopathic carpal tunnel syndrome group was significantly higher than that in controls (odds ratio, 15.8; 95% confidence interval, 3.3-75.7), and age and male sex were independent risk factors for transthyretin amyloid deposition. Our results demonstrate that wild-type transthyretin deposition is a common cause of carpal tunnel syndrome in elderly men. It is likely that many patients develop carpal tunnel syndrome as an initial symptom of senile systemic amyloidosis.ArticleHUMAN PATHOLOGY. 42(11):1785-1791 (2011)journal articl

Targeted adaptive long-read sequencing for discovery of complex phased variants in inherited retinal disease patients

Abstract Inherited retinal degenerations (IRDs) are a heterogeneous group of predominantly monogenic disorders with over 300 causative genes identified. Short-read exome sequencing is commonly used to genotypically diagnose patients with clinical features of IRDs, however, in up to 30% of patients with autosomal recessive IRDs, one or no disease-causing variants are identified. Furthermore, chromosomal maps cannot be reconstructed for allelic variant discovery with short-reads. Long-read genome sequencing can provide complete coverage of disease loci and a targeted approach can focus sequencing bandwidth to a genomic region of interest to provide increased depth and haplotype reconstruction to uncover cases of missing heritability. We demonstrate that targeted adaptive long-read sequencing on the Oxford Nanopore Technologies (ONT) platform of the USH2A gene from three probands in a family with the most common cause of the syndromic IRD, Usher Syndrome, resulted in greater than 12-fold target gene sequencing enrichment on average. This focused depth of sequencing allowed for haplotype reconstruction and phased variant identification. We further show that variants obtained from the haplotype-aware genotyping pipeline can be heuristically ranked to focus on potential pathogenic candidates without a priori knowledge of the disease-causing variants. Moreover, consideration of the variants unique to targeted long-read sequencing that are not covered by short-read technology demonstrated higher precision and F1 scores for variant discovery by long-read sequencing. This work establishes that targeted adaptive long-read sequencing can generate targeted, chromosome-phased data sets for identification of coding and non-coding disease-causing alleles in IRDs and can be applicable to other Mendelian diseases