Pilot detection study of alpha1 antitrypsin deficiency in a targeted population

Screenings for the genetic disorder alpha₁ antitrypsin deficiency (AAT Deficiency) have been one of two models: large screenings of general populations and small targeted detection programs in high‐risk groups. The most appropriate screening and detection methodologies in terms of target populations, subject participation and yield of positive tests, however, have not been well defined. The major objective of this pilot study was to evaluate the effectiveness in terms of participation of two different AAT Deficiency detection programs using a self‐administered fingerstick blood test. Individuals ages 30–60 under the care of a pulmonary physician and with a diagnosis of emphysema, COPD, chronic bronchitis, or bronchiectasis were the targeted population. Participants were offered AAT Deficiency testing in the pulmonary physician's office compared with testing offered through mail. Participation (i.e., frequency of subject participation in the detection program) of two different AAT Deficiency detection programs. Non‐participation was due to fear of self‐administered testing and research studies; women were more likely to participate than men. Eligible subjects were significantly more likely to participate when offered testing by their pulmonary physician in‐office (83%) than mail‐only (42%) (P < 0.02). Although self‐administered genetic testing is available, highest participation in AAT Deficiency detection program was found when offered directly by the physician. This finding may have implications for screening and detection of other genetic diseases. Future studies need to evaluate the yield (i.e., frequency of positive tests) of these detection methodologies in highly targeted populations. © 2001 Wiley‐Liss, Inc.     

Molecular characterization and clinical presentation of HKαα and anti‐HKαα alleles in southern Chinese subjects

The HKαα allele is a rearrangement occurring in the α‐globin gene cluster containing both the ‐α^3.7 and ααα^anti4.2 unequal crossover junctions. The anti‐HKαα allele is the reciprocal product containing both the ‐α^4.2 and ααα^anti3.7 unequal crossover junctions, which had been predicted but had not been detected previously. The phenotypic feature and population frequency of these two unusual alleles were not described. We report the identification of nine individuals carrying the HKαα allele and two individuals carrying the anti‐HKαα allele in southern China and describe their phenotype and haplotype data. The molecular structures of HKαα allele and anti‐HKαα allele were confirmed by two‐round nested polymerase chain reaction assay. The mechanism of origin of both alleles is related to probably simultaneous double crossover. Heterozygotes of HKαα or anti‐HKαα allele show a normal hematological phenotype. Finally, we report the carrier rates of these both alleles in the Guangxi Zhuang Autonomous Region of southern China, namely, ∼0.07% for the HKαα allele and ∼0.02% for the anti‐HKαα allele.     

Attitudes toward an unsolicited approach in relation to status of genetic disease: Exemplified by α1‐antitrypsin deficiency

Knowledge of a genetic disease in an individual raises the questions of whether and how this information should be communicated to his or her family. The aim of the present study was to provide factual information about attitudes towards an unsolicited approach from a physician regarding genetic counseling within affected families. We performed a questionnaire study among patients with α₁‐antitrypsin deficiency, their examined and unexamined relatives, and a control group of Danish citizens. Of 2,146 subjects, the questionnaires were returned by 1,761 (82%), and 1,609 (75%) wanted to participate. Stepwise logistic regression showed that phenotype/subgroup, having descendants, and being female were significantly related to the approval of an unsolicited approach and the informing of relatives. Provided it was difficult for the index case to inform relatives about their risk and about his/her disease, then a total of 75.8% would not proscribe an unsolicited approach by the physician. Most of those for proscribing an unsolicited approach found that relatives should be informed in advance by the index case. The control group of randomly chosen Danes was the most skeptical towards an unsolicited approach. Most individuals found that genetic risk information should be shared with relatives at‐risk. A flexible information policy by the health care system based on active approach towards relatives is acceptable to 75 to 95% of individuals in order to ensure diffusion of genetic risk information within families segregating for a genetic disease with a modifiable outcome. Am. J. Med. Genet. 94:207–213, 2000. © 2000 Wiley‐Liss, Inc.     

A1ATVar: a relational database of human SERPINA1 gene variants leading to α1‐antitrypsin deficiency and application of the VariVis software

We have developed a relational database of human SERPINA1 gene mutations, leading to α₁‐antitrypsin (AAT) deficiency, called A₁ATVar, which can be accessed over the World Wide Web at www.goldenhelix.org/A1ATVar . Extensive information has been extracted from the literature and converted into a searchable database, including genotype information, clinical phenotype, allelic frequencies for the commonest AAT variant alleles, methods of detection, and references. Mutation summaries are automatically displayed and user‐generated queries can be formulated based on fields in the database. A separate module, linked to the FINDbase database for frequencies of inherited disorders allows the user to access allele frequency information for the three most frequent AAT alleles, namely PiM, PiS, and PiZ. The available experimental protocols to detect AAT variant alleles at the protein and DNA levels have been archived in a searchable format. A visualization tool, called VariVis, has been implemented to combine A₁ATVar variant information with SERPINA1 sequence and annotation data. A direct data submission tool allows registered users to submit data on novel AAT variant alleles as well as experimental protocols to explore SERPINA1 genetic heterogeneity, via a password‐protected interface. Database access is free of charge and there are no registration requirements for querying the data. The A₁ATVar database is the only integrated database on the Internet offering summarized information on AAT allelic variants and could be useful not only for clinical diagnosis and research on AAT deficiency and the SERPINA1 gene, but could also serve as an example for an all‐in‐one solution for locus‐specific database (LSDB) development and curation. Hum Mutat 0,1–6, 2008. © 2008 Wiley‐Liss, Inc.     

Recyclable Vinyl‐Functionalized Polyesters via Chemoselective Organopolymerization of Bifunctional α‐Methylene‐δ‐Valerolactone

α‐Methylene‐δ‐valerolactone (MVL) is a bifunctional monomer comprising of a highly stable six‐membered δ‐valerolactone ring and highly reactive C?C bond. Previously, the vinyl‐addition polymerization (VAP) product, namely P(MVL)_VAP, has been formed exclusively. In this study, this conventional chemoselectivity is reversed, wherein organic catalysts are used to enable the first ring‐opening polymerization (ROP) of MVL, exclusively affording a metal‐free, unsaturated polyester, namely P(MVL)_ROP. This challenging goal is achieved by investigating different catalysts, initiators, and reaction conditions. In addition, the formation of two polymers, namely P(MVL)_ROP and P(MVL)_VAP, is easily regulated by varying the polymerization solvent. The resulting P(MVL)_ROP can be easily post‐functionalized to form crosslinked or sulfurized materials; notably, it can be almost completely converted into its monomer thermochemically.     

Molecular analysis of the Pi*Z allele in patients with liver disease

α1‐Antitrypsin (AAT) is the main protease inhibitor in human plasma. There are more than 75 variants of this protein that differ from each other by their isoelectric point. Most of these alleles cause a reduction in AAT levels; the most common allele is Pi*Z. The main complications related to the Pi*Z allele are obstructive pulmonary disease and liver disease. Some Pi*Z allele carriers present cholestatic jaundice and cirrhosis. The Z type is associated with a secretion defect, which leads to deficiency of AAT and to the formation of intrahepatocytic inclusions in affected subjects. The diagnosis of AAT deficiency can be made by different techniques, including molecular analysis, although the final diagnosis should be done in conjunction with demonstration of the periodic acid–Schiff–positive globules on liver biopsy. In this study, specimens of 29 patients with cryptogenic cirrhosis between age 1 month and 18 years, and of 100 controls were submitted to polymerase chain reaction followed by digestion with TaqI enzyme. Five of the 29 patients had undergone liver transplantation. Three patients were heterozygous for the Pi*Z allele, and two were homozygous (allele frequency = 12.07%; 7/58). Among the controls, who represented the population of Porto Alegre, 1 in 100 individuals was heterozygous for the Pi*Z allele, resulting in an allele frequency of 0.5% (1/200). The high frequency of Pi*Z alleles among the patients indicates the usefulness of AAT molecular testing in children with cholestatic jaundice and cirrhosis. © 2001 Wiley‐Liss, Inc.     

22‐S‐Hydroxycholesterol protects against ethanol‐induced liver injury by blocking the auto/paracrine activation of MCP‐1 mediated by LXRα

Chronic ethanol consumption causes hepatic steatosis and inflammation, which are associated with liver hypoxia. Monocyte chemoattractant protein‐1 (MCP‐1) is a hypoxia response factor that determines recruitment and activation of monocytes to the site of tissue injury. The level of MCP‐1 is elevated in the serum and liver of patients with alcoholic liver disease (ALD); however, the molecular details regarding the regulation of MCP‐1 expression are not yet understood completely. Here, we show the role of liver X receptor α (LXRα) in the regulation of MCP‐1 expression during the development of ethanol‐induced fatty liver injury, using an antagonist, 22‐S‐hydroxycholesterol (22‐S‐HC). First, administration of 22‐S‐HC attenuated the signs of liver injury with decreased levels of MCP‐1 and its receptor CCR2 in ethanol‐fed mice. Second, hypoxic conditions or treatment with the LXRα agonist GW3965 significantly induced the expression of MCP‐1, which was completely blocked by treatment with 22‐S‐HC or infection by shLXRα lentivirus in the primary hepatocytes. Third, over‐expression of LXRα or GW3965 treatment increased MCP‐1 promoter activity by increasing the binding of hypoxia‐inducible factor‐1α to the hypoxia response elements, together with LXRα. Finally, treatment with recombinant MCP‐1 increased the level of expression of LXRα and LXRα‐dependent lipid droplet accumulation in both hepatocytes and Kupffer cells. These data show that LXRα and its ligand‐induced up‐regulation of MCP‐1 and MCP‐1‐induced LXRα‐dependent lipogenesis play a key role in the autocrine and paracrine activation of MCP‐1 in the pathogenesis of alcoholic fatty liver disease, and that this activation may provide a promising new target for ALD therapy.Copyright © 2014 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.     

Islets of Langerhans from prohormone convertase‐2 knockout mice show α‐cell hyperplasia and tumorigenesis with elevated α‐cell neogenesis

Antagonism of the effects of glucagon as an adjunct therapy with other glucose‐lowering drugs in the chronic treatment of diabetes has been suggested to aggressively control blood glucose levels. Antagonism of glucagon effects, by targeting glucagon secretion or disabling the glucagon receptor, is associated with α‐cell hyperplasia. We evaluated the influence of total glucagon withdrawal on islets of Langerhans using prohormone convertase‐2 knockout mice (PC2‐ko), in which α‐cell hyperplasia is present from a young age and persists throughout life, in order to understand whether or not sustained glucagon deficit would lead to islet tumorigenesis. PC2‐ko and wild‐type (WT) mice were maintained drug‐free, and cohorts of these groups sampled at 3, 12 and 18 months for plasma biochemical and morphological (histological, immunohistochemical, electron microscopical and image analytical) assessments. WT mice showed no islet tumours up to termination of the study, but PC2‐ko animals displayed marked changes in islet morphology from α‐cell hypertrophy/hyperplasia/atypical hyperplasia, to adenomas and carcinomas, these latter being first encountered at 6–8 months. Islet hyperplasias and tumours primarily consisted of α‐cells associated to varying degrees with other islet endocrine cell types. In addition to substantial increases in islet neoplasia, increased α‐cell neogenesis associated primarily with pancreatic duct(ule)s was present. We conclude that absolute blockade of the glucagon signal results in tumorigenesis and that the PC2‐ko mouse represents a valuable model for investigation of islet tumours and pancreatic ductal neogenesis.     

Hypoxia inducible factor‐1α‐induced interleukin‐33 expression in intestinal epithelia contributes to mucosal homeostasis in inflammatory bowel disease

Intestinal epithelial cells (IECs), an important barrier to gut microbiota, are subject to low oxygen tension, particularly during intestinal inflammation. Hypoxia inducible factor‐1α (HIF‐1α) is expressed highly in the inflamed mucosa of inflammatory bowel disease (IBD) and functions as a key regulator in maintenance of intestinal homeostasis. However, how IEC‐derived HIF‐1α regulates intestinal immune responses in IBD is still not understood completely. We report here that the expression of HIF‐1α and IL‐33 was increased significantly in the inflamed mucosa of IBD patients as well as mice with colitis induced by dextran sulphate sodium (DSS). The levels of interleukin (IL)?33 were correlated positively with that of HIF‐1α. A HIF‐1α‐interacting element was identified in the promoter region of IL‐33, indicating that HIF‐1α activity regulates IL‐33 expression. Furthermore, tumour necrosis factor (TNF) facilitated the HIF‐1α‐dependent IL‐33 expression in IEC. Our data thus demonstrate that HIF‐1α‐dependent IL‐33 in IEC functions as a regulatory cytokine in inflamed mucosa of IBD, thereby regulating the intestinal inflammation and maintaining mucosal homeostasis.     

Involvement of α‐ and β‐catenins and E‐cadherin in the development of mammary phyllodes tumours

Tsang J Y S, Mendoza P, Lam C C F, Yu A M C, Putti T C, Karim R Z, Scolyer R A, Lee C S, Tan P H & Tse G M
(2012) Histopathology  61, 667–674 Involvement of α‐ and β‐catenins and E‐cadherin in the development of mammary phyllodes tumours Aims: Phyllodes tumours (PT) are rare but clinically important fibroepithelial tumours of the breast. β‐Catenin, a key component in Wnt signalling, has been shown to be important in the development of PT. It also functions as a component of the cadherin complex, which may therefore be implicated in PT pathogenesis. By assessing stromal α‐catenin, β‐catenin and E‐cadherin expression in 158 PT cases using immunohistochemistry and examining associations with clinicopathological features, we aimed to determine the role of these proteins in PT pathogenesis. Methods and results: Cytoplasmic β‐catenin correlated with α‐catenin expression. A significantly higher expression of both markers was observed in borderline than in benign PT (P = 0.003 and <0.001, respectively), but a lower level was found in malignant PT. Cytoplasmic E‐cadherin expression was significantly higher in borderline and malignant than in benign PT (P = 0.001 and 0.012, respectively), but was not correlated with other markers. Both E‐cadherin and α‐catenin showed stronger correlations with histological parameters than β‐catenin. α‐Catenin showed a significant correlation with recurrence (P = 0.005 and 0.016, respectively). Conclusions: α‐ and β‐catenins may be important in the early stages of PT development, while E‐cadherin may be required for malignant development. The correlation of α‐catenin expression with tumour recurrence may be relevant in predicting PT behaviour.     

The stimulatory effect of α‐melanotropin on progesterone release from rat granulosa cells is inhibited by interleukin‐1β and by tumour necrosis factor‐α

Aim: Several studies have shown that a variety of peptides and cytokines are involved in ovarian regulatory mechanisms; however, their exact function is still unclear. In this work we study whether the administration of peptide α‐melanotropin and the cytokines interleukin‐1β (IL‐1β) and tumour necrosis factor‐α (TNF‐α) on their own modify the release of progesterone in cultured granulosa cells (GC) from pro‐oestrous rats. We also investigate an interaction between these cytokines and α‐melanotropin in the modulation of progesterone secretion. Methods: Granulosa cells were collected from the ovaries of female Wistar rats and cultured for up to 24 h in the presence of different concentrations of α‐melanotropin, cytokines or a combination of both. Progesterone concentration was measured by radioimmunoassay. Results: The addition of α‐melanotropin in a dose of 0.01 and 0.1 mm had no effect on progesterone release, whereas a dose of 1 mm significantly increased progesterone release (P < 0.01) compared with the control culture. Progesterone release was not modified when different concentrations of interleukin‐1β or TNF‐α were added to the cell cultures. However, when interleukin‐1β or TNF‐α were added simultaneously with 1 μm α‐melanotropin, a significant reduction (P < 0.01 for interleukin‐1β and P < 0.05 for TNF‐α) of the steroid release was found with respect to the α‐melanotropin‐treated group. Conclusions: These results lead us to suggest that, although α‐melanotropin stimulates progesterone release in pre‐ovulatory GC, this effect is blocked by the presence of interleukin‐1β or TNF‐α.     

Genetic association study between α 1‐antichymotrypsin polymorphism and Alzheimer disease in Chinese Han population

We investigated a common signal peptide polymorphism in the α 1‐antichymotrypsin (ACT) gene in 125 sporadic Alzheimer disease (AD) patients and 141 healthy control subjects in Chinese Han population. We found no significant difference in the distribution of ACT polymorphism between AD cases and controls, and failed to detect any effects of ACT genotypes associated with AD. Thus, our data do not support the involvement of ACT polymorphism in the risk of AD in Chinese Han population. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:133–135, 2000. © 2000 Wiley‐Liss, Inc.     

Secreted IL‐1α promotes T‐cell activation and expansion of CD11b+Gr1+ cells in carbon tetrachloride‐induced liver injury in mice

Interleukin‐1α is mainly expressed on the cell membrane, but can also be secreted during inflammation. The roles of secreted and membrane IL‐1α in acute liver inflammation are still not known. Here, we examined the functions of secreted and membrane IL‐1α in a mouse model of carbon tetrachloride‐induced acute liver injury. We show that secreted IL‐1α aggravates liver damage and membrane IL‐1α slightly protects mice from liver injury. Further studies showed that secreted IL‐1α promotes T‐cell activation. It also increased the expansion of CD11b⁺Gr1⁺ myeloid cells, which may serve as a negative regulator of acute liver inflammation. Moreover, secreted IL‐1α induced IL‐6 production from hepatocytes. IL‐6 neutralization reduced the proliferation of CD11b⁺Gr1⁺ myeloid cells in vivo. CCL2 and CXCL5 expression was increased by secreted IL‐1α in vitro and in vivo. Antagonists of the chemokine receptors for CCL2 and CXCL5 significantly reduced the migration of CD11b⁺Gr1⁺ myeloid cells. These results demonstrate that secreted and membrane IL‐1α play different roles in acute liver injury. Secreted IL‐1α could promote T‐cell activation and the recruitment and expansion of CD11b⁺Gr1⁺ myeloid cells through induction of CCL2, CXCL5, and IL‐6. The controlled release of IL‐1α could be a critical regulator during acute liver inflammation.     

Human mesenchymal stromal cells modulate T‐cell responses through TNF‐α‐mediated activation of NF‐κB

Although mesenchymal stromal cells (MSCs) possess the capacity to modulate immune responses, little is known about the mechanisms that underpin these processes. In this study, we show that immunosupression is mediated by activation of nuclear factor kappa B (NF‐κB) in human MSCs. This pathway is activated by TNF‐α that is generated following TCR stimulation of T cells. Inhibition of NF‐κB through silencing of IκB kinase β or the TNF‐α receptor abolishes the immunosuppressive capacity of MSCs. Our data also indicate that MSC‐associated NF‐κB activation primarily leads to inhibition of T‐cell proliferation with little effect on expression of the activation markers CD69 and CD25. Thus, our data support the hypothesis that the TNF‐α/NF‐κB signalling pathway is required for the initial priming of immunosuppressive function in human MSCs. Interestingly, drugs that interfere with NF‐κB activation significantly antagonise the immunoregulatory effect of MSCs, which could have important implications for immunosuppression regimens in the clinic.     

JNK1 and JNK2 regulate α‐SMA in hepatic stellate cells during CCl4‐induced fibrosis in the rat liver

Following liver injuries, hepatic stellate cells (HSCs) express α‐SMA. Mitogen activated protein kinase (MAPK) signaling pathways mediate α‐SMA expression in distinct cell types. However, the regulation of α‐SMA expression by MAPKs in HSCs has been rarely studied. We aimed to study the role of MAPKs in the activation of HSCs during liver fibrosis. Liver fibrosis of rats was induced by carbon tetrachloride. HSC‐T6 cells, murine embryonic fibroblasts, JNK1^?/? and JNK2^?/? cells were used for in vitro studies. Immunohistochemistry and immunoblot analysis were used. We have found that the expression of JNK and α‐SMA co‐localized in HSCs during liver fibrosis, but ERK and p38 expressed in macrophages. The expression of α‐SMA was up‐regulated by JNK1 and JNK2 in non‐stress condition. Under TGF‐β stimulation, however, the level α‐SMA expression was increased by only JNK1, but not significantly changed by JNK2. We suggest that JNKs are responsible for α‐SMA regulation, and especially JNK1 has a major role in up‐regulation of α‐SMA expression in HSCs under stress condition induced by TGF‐β during liver fibrosis.