[Clinical phenotype, genetic characteristics, and creation of immortalized cell lines for patients from a pedigree affected with Hunter syndrome].

OBJECTIVE: To explore the clinical phenotype and genetic variant in a Chinese pedigree affected with Hunter syndrome and create immortalized cell lines for the affected pedigree members. METHODS: A pedigree of six members who had visited Xi'an Children's Hospital in July 2022 was selected as the study subject. Clinical data was collected. Whole exome sequencing was carried out for the pedigree members. Candidate variant was verified by Sanger sequencing. In addition, peripheral B lymphocytes were transfected with Epstein-Barr virus to create immortalized cell lines, which were then subjected to enzyme activity analysis. RESULTS: The patient, a five-year-and-seven-month-old boy, had exhibited stiff limbs and enlarged joints. He had developed hernia, scaphocephaly, and barrel chest from 3 months of age. His uncle also had stiff limbs, poor hearing, blindness, and right oblique inguinal hernia. Above features had resembled those of Hunter syndrome. Genetic testing revealed that both the child and his uncle had harbored an IDS (NM_000202.8): c.823G>A (p.D275N) variant, which was unreported previously. Bioinformatic analysis indicated that the D275 to be a highly conserved site, and the D275N variant may affect the stability of the protein's spatial conformation, thereby decrease the catalytic activity of the enzyme. The successfully constructed immortalized lymphoblastoid cell lines for the child and his parents showed increased volume, irregular shape, burr structure and cluster growth. And the value of IDS activity of the patient's immortalized lymphoblastoid cells was below the limit of detection. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as likely pathogenic (PS3+PM2_Supporting+PM5+PP1+PP3). CONCLUSION: Above finding has enriched the phenotypic and mutational spectra of Hunter syndrome, and provided a basis for the genetic counseling for this pedigree. The creation of immortalized cell lines has offered a model for further investigation of the impact of variant on the function of IDS and development of targeted drugs.

Tamoxifen induces the development of hernia in mice by activating MMP-2 and MMP-13 expression.

Hernia is a disease with defects in collagen synthesis/metabolism. However, the underlying mechanisms for hernia formation have not been fully defined. Tamoxifen is a selective estrogen receptor modulator and used for patients with breast cancer. Tamoxifen also has pleiotropic and side effects. Herein, we report that tamoxifen treatment resulted in an appearance of a large bulge in the low abdomen between the hind legs in male but not in female mice. The autopsy demonstrated that the low abdominal wall was broken and a large amount of intestine herniated out of the abdominal cavity. Histological analysis indicated that tamoxifen caused structural abnormalities in the low abdominal wall which were associated with decreased type II collagen content. Furthermore, we determined increased matrix metalloproteinase-2 (MMP-2) and MMP-13 expression in the tissue. In vitro, tamoxifen induced MMP-2 and MMP-13 expression in fibroblasts. The promoter activity analysis and ChIP assay demonstrate that induction of MMP-13 expression was associated with activation of JNK-AP-1 and ERK1/2 signaling pathways while induction of MMP-2 expression was related to activation of the ERK1/2 signaling pathway. Taken together, our study establishes a novel murine hernia model, defines a severe side effect of tamoxifen, and suggests a caution to male patients receiving tamoxifen treatment.

On the use of the transmission disequilibrium test to detect pseudo-autosomal variants affecting traits with sex-limited expression.

We herein describe the realization of a genome-wide association study for scrotal hernia and cryptorchidism in Norwegian and Belgian commercial pig populations. We have used the transmission disequilibrium test to avoid spurious associations due to population stratification. By doing so, we obtained genome-wide significant signals for both diseases with SNPs located in the pseudo-autosomal region in the vicinity of the pseudo-autosomal boundary. By further analyzing these signals, we demonstrate that the observed transmission disequilibria are artifactual. We determine that transmission bias at pseudo-autosomal markers will occur (i) when analyzing traits with sex-limited expression and (ii) when the allelic frequencies at the marker locus differ between X and Y chromosomes. We show that the bias is due to the fact that (i) sires will preferentially transmit the allele enriched on the Y (respectively X) chromosome to affected sons (respectively daughters) and (ii) dams will appear to preferentially transmit the allele enriched on the Y (respectively X) to affected sons (respectively daughters), as offspring inheriting the other allele are more likely to be non-informative. We define the conditions to mitigate these issues, namely by (i) extracting information from maternal meiosis only and (ii) ignoring trios for which sire and dam have the same heterozygous genotype. We show that by applying these rules to scrotal hernia and cryptorchidism, the pseudo-autosomal signals disappear, confirming their spurious nature.

Absence of chromosomal abnormalities in herniated orbital fat.

AIMS: Lipomatous tumours of the orbit are rare, and can sometimes be difficult to characterize. Herniated orbital fat is thought to be a reactive process, but its presentation can mimic a lipomatous tumour such as an atypical lipomatous tumour or spindle cell/pleomorphic lipoma. Genetic studies to determine if it is indeed a reactive process rather than an adipocytic neoplasm have not been performed. METHODS AND RESULTS: Four samples of herniated orbital fat were reviewed clinically, histopathologically and immunohistochemically. Array comparative genomic hybridization (aCGH) was used to search for genome-wide copy number alterations within the tumours. Histological evaluation revealed that all four tumours contained collections of adipocytes surrounded by fibrous septae. Lochkern cells and floret-like multinucleated giant cells were present, consistent with herniated orbital fat. CD34 was positive in all tumours. Staining for MDM2 and CDK4 was negative. ACGH analysis demonstrated no copy number alterations. CONCLUSIONS: Herniated orbital fat may share some histopathological features with lipoma and atypical lipomatous tumour, but the absence of copy number gains or losses is consistent with the impression that herniated orbital fat is a reactive process. Genetic analysis may be another method to help differentiate herniated orbital fat from a lipomatous orbital tumour when the diagnosis is in question.

Comparison of protein immunoprecipitation-multiple reaction monitoring with ELISA for assay of biomarker candidates in plasma.

Quantitative analysis of protein biomarkers in plasma is typically done by ELISA, but this method is limited by the availability of high-quality antibodies. An alternative approach is protein immunoprecipitation combined with multiple reaction monitoring mass spectrometry (IP-MRM). We compared IP-MRM to ELISA for the analysis of six colon cancer biomarker candidates (metalloproteinase inhibitor 1 (TIMP1), cartilage oligomeric matrix protein (COMP), thrombospondin-2 (THBS2), endoglin (ENG), mesothelin (MSLN) and matrix metalloproteinase-9 (MMP9)) in plasma from colon cancer patients and noncancer controls. Proteins were analyzed by multiplex immunoprecipitation from plasma with the ELISA capture antibodies, further purified by SDS-PAGE, digested and analyzed by stable isotope dilution MRM. IP-MRM provided linear responses (r = 0.978-0.995) between 10 and 640 ng/mL for the target proteins spiked into a "mock plasma" matrix consisting of 60 mg/mL bovine serum albumin. Measurement variation (coefficient of variation at the limit of detection) for IP-MRM assays ranged from 2.3 to 19%, which was similar to variation for ELISAs of the same samples. IP-MRM and ELISA measurements for all target proteins except ENG were highly correlated (r = 0.67-0.97). IP-MRM with high-quality capture antibodies thus provides an effective alternative method to ELISA for protein quantitation in biological fluids.

A Potential miRNA-mRNA Network for Dementia and Hernia Crosstalk.

BACKGROUND: It has been reported that there may be a potential link between hernia and dementia. However, the exact mechanisms of their association have not been established. This study is aimed at constructing miRNA-mRNA networks to elucidate on the potential link between dementia and hernia. METHODS: Gene expression profiles for dementia, herniation, and skeletal muscle were downloaded from the GEO database after which differentially expressed mRNAs and miRNAs were obtained. In addition, fascia tissue samples were obtained during surgery. A total of 41 patients were recruited in this study, and expression levels of candidate genes were examined using quantitative RT-PCR. Luciferase reporter gene assays were used to identify potential miRNA-mRNA regulatory pathways. RESULTS: Differentially expressed mRNAs and miRNAs were screened. A potential miRNA-mRNA network revealing the crosstalk mechanism between herniation and dementia was identified. Single cell analysis revealed that PI16 was highly enriched in adipose tissues, skeletal muscles, and in the skin. GSEA enrichment analysis showed that PI16 is involved in adipose metabolism, muscle functions, and energy metabolism. In clinical samples, PI16 was found to be upregulated in hernia, while miR-4451 was found to be downregulated. The luciferase reporter gene assay revealed that downregulation of circulating miR-4451 may be responsible for the upregulated PI16 expression in hernia sacs. CONCLUSIONS: We constructed an miRNA-mRNA network that shows the potential association between dementia and hernia. We also found that miR-4451 regulates the PI16 expression, which may be a key target and biomarker for hernia pathogenesis and dementia crosstalk.

Superior herniation of the thymus into the neck--a familial pattern.

Superior herniation of normal mediastinal thymus into the anterior neck is a rare cause of neck masses in children. It is defined as intermittent migration of the broadest part of the normal thymus out of the thorax into the suprasternal region during Valsalva maneuver with an increase in the intrathoracic pressure. The fact that the mass apparent only during Valsalva maneuver and typical ultrasound characteristics usually allow the diagnosis but computerized tomography scan or magnetic resonance imaging is necessary to assess the extent of the mass. We report the first and the only siblings with the most dramatic degree of superior herniation of normal mediastinal thymus. We discuss the findings of imaging and the differential diagnosis. We try to remind this entity to avoid unnecessary biopsy or surgery and their potential risk of altering immune function.

Identification of endogenous normalizing genes for expression studies in inguinal ring tissue for scrotal hernias in pigs.

The use of reference genes is required for relative quantification in gene expression analysis and since the stability of these genes could be variable depending on the experimental design, it has become indispensable to test the reliability of endogenous genes. Therefore, this study evaluated 10 reference candidate genes in two different experimental conditions in order to obtain stable genes to be used as reference in expression studies related to scrotal hernias in pigs. Two independent experiments were performed: one with 30 days-old MS115 pigs and the other with 60 days-old Landrace pigs. The inguinal ring/canal was collected, frozen and further submitted to real-time PCR analysis (qPCR). For the reference genes stability evaluation, four tools were used: GeNorm in the SLqPCR, BestKeeper, NormFinder and Comparative CT. A general ranking was generated using the BruteAggreg function of R environment. In this study, the RPL19 was one of the most reliable endogenous genes for both experiments. The breed/age effects influenced the expression stability of candidate reference genes evaluated in the inguinal ring of pigs. Therefore, this study reinforces the importance of evaluating the stability of several endogenous genes previous their use, since a consensual set of reference genes is not easily obtained. Here, two sets of genes are recommended: RPL19, RPL32 and H3F3A for 30-days MS115 and PPIA and RPL19 for the 60 days-old Landrace pigs. This is the first study using the inguinal ring tissue and the results can be useful as an indicative for other studies working with gene expression in this tissue.

Genome-wide linkage analysis of inguinal hernia in pigs using affected sib pairs.

BACKGROUND: Inguinal and scrotal hernias are of great concern to pig producers, and lead to poor animal welfare and severe economic loss. Selection against these conditions is highly preferable, but at this time no gene, Quantitative Trait Loci (QTL), or mode of inheritance has been identified in pigs or in any other species. Therefore, a complete genome scan was performed in order to identify genomic regions affecting inguinal and scrotal hernias in pigs. Records from seedstock breeding farms were collected. No clinical examinations were executed on the pigs and there was therefore no distinction between inguinal and scrotal hernias. The genome scan utilised affected sib pairs (ASP), and the data was analysed using both an ASP test based on Non-parametric Linkage (NPL) analysis, and a Transmission Disequilibrium Test (TDT). RESULTS: Significant QTLs (p < 0.01) were detected on 8 out of 19 porcine chromosomes. The most promising QTLs, however, were detected in SSC1, SSC2, SSC5, SSC6, SSC15, SSC17 and SSCX; all of these regions showed either statistical significance with both statistical methods, or convincing significance with one of the methods. Haplotypes from these suggestive QTL regions were constructed and analysed with TDT. Of these, six different haplotypes were found to be differently transmitted (p < 0.01) to healthy and affected pigs. The most interesting result was one haplotype on SSC5 that was found to be transmitted to hernia pigs with four times higher frequency than to healthy pigs (p < 0.00005). CONCLUSION: For the first time in any species, a genome scan has revealed suggestive QTLs for inguinal and scrotal hernias. While this study permitted the detection of chromosomal regions only, it is interesting to note that several promising candidate genes, including INSL3, MIS, and CGRP, are located within the highly significant QTL regions. Further studies are required in order to narrow down the suggestive QTL regions, investigate the candidate genes, and to confirm the suggestive QTLs in other populations. The haplotype associated with inguinal and scrotal hernias may help in achieving selection against the disorder.

Hernia disease and collagen gene regulation: are there clues for intervention?

Collagens belong to the most abundant proteins in the body. After tissue injury, a coordinated regulation of collagen gene expression guides the formation of a provisional matrix that subsequently evolves into a mature scar with tensile strength. In the following, knowledge regarding collagen gene regulation that may provide insight into how to specifically address the biological problem of soft tissue weakness and recurrent hernia disease is summarized.

Genetic analysis of the cause of gastroschisis in the HLG mouse strain.

An inbred mouse strain HLG shows a high incidence of gastroschisis after X-ray exposure to the zygotes. About 11% of the fetuses display this malformation after irradiation with 1 Gy. The C57BL-strain does not show the increased frequency of gastroschisis after radiation-exposure to the zygotes. The genetic background of this malformation was investigated in a backcross of HLG x C57BL females to HLG males. The pregnant HLG x C57BL females were irradiated in a stage in which the (HLG x C57BL) x HLG [BC1] embryos were in the 1-cell stage. The frequency of gastroschisis in the BC1 generation was compared with a genetic model of a single recessive mutation with 11% penetrance. This frequency does not fit a single-locus inheritance. The number of loci involved was estimated to be about two or three. HLG mouse strain may be a valuable animal model in the study of polygenic traits.

[Familial cervical lung hernia: a report of 4 cases in a family].

In this study 4 members of three generations in one family suffered cervical lung hernia was reported. In this 4 cases, there were 2 males, and 2 females, and 3 cases found on the right side, 1 on the left side of neck. Their ages were 55, 32, 5 and 6, respectively. X-ray examinations and CT scan showed typical characteristics. Chromosome examination analysis with high resolution G band was done for 2 of them, no abnormal changes were found. From pedigree analysis we think this type of disease is an autosomal dominant hereditary disease. This familial abnormality has not been found in medical literature, and so we suggest a namelity "Familial cervical lung hernia".

Lack of fibulin-3 causes early aging and herniation, but not macular degeneration in mice.

A mutation in the EFEMP1 gene causes Malattia Leventinese, an inherited macular degenerative disease with strong similarities to age-related macular degeneration. EFEMP1 encodes fibulin-3, an extracellular matrix protein of unknown function. To investigate its biological role, the murine Efemp1 gene was inactivated through targeted disruption. Efemp1(-/-) mice exhibited reduced reproductivity, and displayed an early onset of aging-associated phenotypes including reduced lifespan, decreased body mass, lordokyphosis, reduced hair growth, and generalized fat, muscle and organ atrophy. However, these mice appeared to have normal wound healing ability. Efemp1(-/-) mice on a C57BL/6 genetic background developed multiple large hernias including inguinal hernias, pelvic prolapse and protrusions of the xiphoid process. In contrast, Efemp1(-/-) mice on a BALB/c background rarely had any forms of hernias, indicating the presence of modifiers for fibulin-3's function in different mouse strains. Histological analysis revealed a marked reduction of elastic fibers in fascia, a thin layer of connective tissue maintaining and protecting structures throughout the body. No apparent macular degeneration associated defects were found in Efemp1(-/-) mice, suggesting that loss of fibulin-3 function is not the mechanism by which the mutation in EFEMP1 causes macular degeneration. These data demonstrate that fibulin-3 plays an important role in maintaining the integrity of fascia connective tissues and regulates aging.

Familial synspondylism: progressive scoliosis and multiple hernias in a kinship.

A new genetic syndrome is reported of congenital lordoscoliosis due to lumbar segmentation defects and incomplete formation of lumbar vertebrae. The defect arose as a spontaneous mutation and was transmitted in an autosomal dominant fashion. The kindred included a mother and her three offspring. These affected individuals had several dysmorphic features including cavus feet and micrognathia. In addition the syndrome was associated with multiple hernias including inguinal, ventral, and diaphragmatic. These associated problems led to the early death of the first child at age 7 months. The lumbar scoliosis was already evident by that time. The progressive nature of the scoliosis was documented, especially in one child who was lost to follow-up and who was initially seen with a severe spinal deformity. Surgical management was required in members of the kindred, but because of differences in age and severity at the time of surgery, the techniques varied.

Necropsy findings in a fetus with a 46,XY,dic t(X;21)(p11.1;p11.1).

We report the findings in a fetus terminated because of multiple abnormalities diagnosed on ultrasound, including asymmetry of the limbs, a hypoplastic diaphragm, unilateral duplex kidney with a double ureter, unilateral cystic kidney, and congenital heart disease including total pulmonary atresia. Cytogenetic studies showed an unbalanced translocation of the long arm of the X chromosome to chromosome 21, resulting in a 46,XY,dic t(X;21)(p11.1;p11.1) karyotype. The cytogenetics were confirmed by non-isotopic in situ hybridisation using probes specific to pericentric alphoid repeats. Parental chromosomes were normal indicating this to be a de novo translocation. It is suggested that the inactivation of the long arm of the X chromosome has resulted in an effective monosomy for chromosome 21.

Magnetic resonance imaging of hereditary hernias of the peroneus longus muscle.

Herniation of the left peroneus longus muscle was present in three male members of the same family, being the first reported case of this condition in a familial setting. The hernias were differentiated from other mass lesions and varices by magnetic resonance imaging. The images demonstrated a fascial defect originating in the area where vessels and nerves penetrate the fascia, suggesting that the three men had a congenital weakness in the fascia.

Dubowitz syndrome.

Four children including two siblings with Dubowitz syndrome are presented. All four were preterm or small-for-dates. On the basis of their symptoms, it is suggested that infantile eczema is not an essential sign of the disorder, whereas the high frequency of hernia, strabism and upward slant of the palpebral fissures is underestimated in the literature.