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The SERPINA1 gene encodes the serine protease inhibitor alpha-1 antitrypsin (AAT) and is located on chromosome 14q31-32.3 in a cluster of homologous genes likely formed by exon duplication. AAT has a variety of anti-inflammatory properties. Its clinical relevance is best illustrated by the genetic disease alpha-1 antitrypsin deficiency (AATD) which is associated with an increased risk for chronic obstructive pulmonary disease (COPD) and cirrhosis. While 2 single nucleotide polymorphisms (SNPs) , S and Z, are responsible for more than 95% of all individuals with AATD, there are a number of rare variants associated with deficiency and dysfunction, as well as those associated with normal levels and function. Our laboratory has identified a number of novel AAT alleles that we report in this manuscript. We screened more than 500,000 individuals for AATD alleles through our testing program over the past 20 years. The characterization of these alleles was accomplished by DNA sequencing, measurement of AAT plasma levels and isoelectric focusing at pH 4-5. We report 22 novel AAT alleles discovered through our screening programs, such as Z(little rock) and QO(chillicothe), and review the current literature of known AAT genetic variants.

This study aimed to identify if individuals with mild to severe alpha-1 antitrypsin deficiency (AATD) are at higher risk for developing obstructive sleep apnea (OSA) than the general population. A seven-question sleep apnea risk assessment questionnaire, STOP-BAG, was applied to 2338 participant responses from the Alpha-1 Coded Testing Study (ACT) and 4638 participant responses from the Kentucky Behavioral Risk Factor Survey (KyBRFS). Propensity scores were generated from a logistic regression model using continuous variables of age and body mass index (BMI). STOP-BAG scores were analyzed using chi-square analysis on this matched cohort to assess OSA risk in AATD. Self-reported OSA was higher in the KyBRFS cohort (14.5%) than in individuals with mild or severe AATD (11.2%) (p = 0.012). However, a higher percentage of the AATD cohort met clinically meaningful thresholds for STOP-BAG scores ≥ 5 (22.7%) than the KyBRFS cohort (13.0%) (p = 0.001). These differences persisted despite 1:1 propensity score matching on age and BMI to account for differences in baseline characteristics. No statistically significant difference in OSA risk between AATD genotypes was found. AATD appears to have higher risk for OSA than the general population. The 11.2% prevalence of diagnosed OSA in the AATD population is much lower than symptom scores would predict. Further studies are needed to validate the possibility that elastin loss is involved in OSA pathogenesis.

There are a number of respiratory diseases characterized by the presence of excess neutrophil elastase (NE) activity in tissues, including cystic fibrosis and chronic obstructive pulmonary disease (COPD). NE is considered a primary contributor to disease development, but the precise mechanism has yet to be fully determined. We hypothesized that NE alters the function of macrophages (Mɸ) which play a critical role in many physiological processes in healthy lungs. We demonstrate that monocyte-derived Mɸ exposed to NE releases active matrix metalloproteinases (MMPs), increase expression of pro-inflammatory cytokines TNFα, IL-1β, and IL-8, and reduce capacity to phagocytose bacteria. Changes in Mɸ function following NE treatment were accompanied by increased adhesion and cytoskeleton re-arrangement, indicating the possibility of integrin involvement. To support this observation, we demonstrate that NE induces phosphorylation of kinases from the Src kinase family, a hallmark of integrin signaling activation. Moreover, pretreatment of Mɸ with a specific Src kinase inhibitor, PP2 completely prevents NE-induced pro-inflammatory cytokine production. Taken together these findings indicate that NE participates in lung destruction not only through direct proteolytic degradation of matrix proteins, but also through activation of Mɸ inflammatory and proteolytic functions.

Background Most patients with alpha-1 antitrypsin deficiency remain undiagnosed and therefore do not benefit from current therapies or become eligible for research studies of new treatments under development. Improving the detection rate for AATD is therefore a high priority for the Alpha-1 Foundation. A workshop was held on June 23, 2019 in Orlando, Florida during which stakeholders from the research, pharmaceutical, and patient communities focused on the topic of alpha-1 antitrypsin deficiency detection. Results A variety of detection strategies have been explored in the past and new approaches are emerging as technology advances. Targeted detection includes patients with chronic obstructive pulmonary disease, unexplained chronic liver disease, and family members of affected individuals. Newborn screening, electronic medical record data mining, and direct-to-consumer testing remain options for future detection strategies. Conclusion These meeting proceedings can serve as a basis for innovative approaches to the detection of alpha-1 antitrypsin deficiency.

Background and purpose Pulmonary hypertension (PH) secondary to chronic lung disease (World Health Organization Group 3 PH) is deadly, with lung transplant being the only available long-term treatment option. Myeloid-derived cells are known to affect progression of both pulmonary fibrosis and PH, although the mechanism of action is unknown. Therefore, we investigated the effect of myeloid cell proliferation induced by emergency myelopoiesis on development of PH and therapy directed against programmed death-ligand 1 (PD-L1), expressed by myeloid cells in prevention of pulmonary vascular remodelling. Experimental approach LysM.Cre-DTR ("mDTR") mice were injected with bleomycin (0.018 U·g-1 , i.p.) while receiving either vehicle or diphtheria toxin (DT; 100 ng, i.p.) to induce severe PH. Approximately 4 weeks after initiation of bleomycin protocol, right ventricular pressure measurements were performed and tissue samples collected for histologic assessment. In a separate experiment, DT-treated mice were given anti-PD-L1 antibody (αPD-L1; 500 μg, i.p.) preventive treatment before bleomycin administration. Key results Mice undergoing induction of emergency myelopoiesis displayed more severe PH, right ventricular remodelling and pulmonary vascular muscularization compared to controls, without a change in lung fibrosis. This worsening of PH was associated with increased pulmonary myeloid-derived suppressor cell (MDSC), particularly polymorphonuclear MDSC (PMN-MDSC). Treatment with αPD-L1 normalized pulmonary pressures. PD-L1 expression was likewise found to be elevated on circulating PMN-MDSC from patients with interstitial lung disease and PH. Conclusions and implications PD-L1 is a viable therapeutic target in PH, acting through a signalling axis involving MDSC. Linked articles This article is part of a themed issue on Risk factors, comorbidities, and comedications in cardioprotection. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.1/issuetoc.

Background: Alpha-1 antitrypsin deficiency (AATD) is under-recognized, prompting the need for enhanced detection strategies. The primary aim of this study is to determine the feasibility of using the electronic medical record (EMR) and linked electronic patient messages (EPM) to encourage AATD testing by patients with chronic obstructive pulmonary disease (COPD). Methods: Study participants were eligible, untested adult patients who were prescribed an inhaled medication which is exclusively Food and Drug Administration-approved for treating COPD. Eligible patients received a message with basic information about AATD and availability of free, home-based AATD testing. Through a collaboration with the Alpha-1 Foundation’s Alpha-1 Coded Testing (ACT) study, patients referred to home-based testing through EPM were flagged. The effectiveness of the electronic message was evaluated by the proportion of patients who underwent testing, and the rate of detecting individuals with severe deficiency of AAT among those tested. Results: A total of 12,369 patients on eligible inhalers were screened; 5430 patients met all criteria and received an EPM. During the study, 396 patients (7.3%) fully requested an ACT kit. Of these, 209 patients (52.8%) returned the test sample and received genotyping results; 65.5%, had a normal AAT genotype (PI*MM), 31.6% were heterozygotes for a deficient allele (PI*MS, PI*MZ and PI*M/Null rare), and 2.9% had severe deficiency of alpha-1 antitrypsin (PI*SZ, PI*ZZ, PI*S/Null rare). Conclusion: While the response rate and test return rate were low, the rate of detecting individuals with AATD using this detection strategy exceeds that of many prior strategies. As such, while requiring independent validation in other populations, this detection strategy holds promise.

Pulmonary hypertension complicates the care of many patients with chronic lung diseases (defined as Group 3 pulmonary hypertension), yet the mechanisms that mediate the development of pulmonary vascular disease are not clearly defined. Despite being the most prevalent form of pulmonary hypertension, to date there is no approved treatment for patients with disease. Myeloid-derived suppressor cells (MDSCs) and endothelial cells in the lung express the chemokine receptor CXCR2, implicated in the evolution of both neoplastic and pulmonary vascular remodeling. However, precise cellular contribution to lung disease is unknown. Therefore, we used mice with tissue-specific deletion of CXCR2 to investigate the role of this receptor in Group 3 pulmonary hypertension. Deletion of CXCR2 in myeloid cells attenuated the recruitment of polymorphonuclear MDSCs to the lungs, inhibited vascular remodeling, and protected against pulmonary hypertension. Conversely, loss of CXCR2 in endothelial cells resulted in worsened vascular remodeling, associated with increased MDSC migratory capacity attributable to increased ligand availability, consistent with analyzed patient sample data. Taken together, these data suggest that CXCR2 regulates MDSC activation, informing potential therapeutic application of MDSC-targeted treatments.

Alpha1-antitrypsin (AAT) deficiency predisposes individuals to emphysema and liver diseases such as cirrhosis and hepatocellular carcinoma. The deficiency results from mutations in the SERPIN1A gene encoding AAT molecules that cause hepatotoxic retention within the endoplasmic reticulum. Since the E342K mutation is the basis for destabilization leading to lung and liver pathologies, we used the crystal structure of the mutated AAT as the basis for molecular docking selection of candidate compounds that may bind and stabilize the 342K structural pocket. We identified compounds that inhibited intracellular accumulation of AAT in hepatocytes in vitro. These data suggest that drug binding to a structural site encoded by a mutation associated with AAT deficiency has the potential for clinical utility by modulating conformational transitions.

Background The immunomodulatory effects of statins on vaccine response remain uncertain. Therefore, the objective of this study was to determine if atorvastatin enhances pneumococcal-specific antibody titer following 23-valent pneumococcal polysaccharide vaccination. Methods Double-blind, placebo-controlled, single-center randomized clinical trial entitled StatVax. Subjects were enrolled between June and July 2014 and followed up through September 2014. 33 healthy volunteers signed informed consent after volunteer sampling. 11 participants were excluded; 22 healthy volunteers without prior pneumococcal vaccination were enrolled and completed the study. Participants were randomized to receive a 28-day course of 40 mg atorvastatin (n = 12) or matching lactose placebo (n = 10). On day 7 of treatment, Pneumovax 23 was administered intramuscularly. The primary outcome was fold change in total pneumococcal-specific antibody titer determined by a ratio of post-vaccination titer over baseline titer. Secondary outcomes included serotype-specific pneumococcal antibody titer, seroconversion, complete blood counts (CBC), erythrocyte sedimentation rate (ESR), and serum cytokine analysis. Results Of the 22 randomized patients (mean age, 23.86; SD, 4.121; 11 women [50%]), 22 completed the trial. Total anti-pneumococcal antibody titer in the atorvastatin group went from a baseline mean of 32.58 (SD, 15.96) to 147.7 (SD, 71.52) μg/mL at 21 days post-vaccination while titer in the placebo group went from a mean of 30.81 (SD, 13.04) to 104.4 (SD, 45) μg/mL. When comparing fold change between treatment groups, there was a significant increase in fold change of total anti-pneumococcal antibody titer in the atorvastatin group compared to the placebo group (2-way ANOVA, p = .0177). Conclusions Atorvastatin enhances antigen-specific primary humoral immune response to a T cell-independent pneumonia vaccination. Pending confirmation by larger cohort studies of target populations, peri-vaccination conventional doses of statins can become a novel adjuvant for poorly-immunogenic polysaccharide-based vaccines. Trial Registration: clinicaltrials.gov Identifier: NCT02097589

Background & Aims Alpha-1 antitrypsin deficiency (AATD) is an uncommonly recognized cause of liver disease in adults, with descriptions of its natural history limited to case series and patient-reported data from disease registries. Liver pathology is limited to selected patients or unavailable. Therefore, we aimed to determine the prevalence and severity of liver fibrosis in an adult AATD population who were not known to have cirrhosis, while defining risk factors for fibrosis and testing non-invasive markers of disease. Methods A total of 94 adults with classic genotype ‘PI*ZZ’ AATD were recruited from North America and prospectively enrolled in the study. Liver aminotransferases and markers of synthetic function, transient elastography, and liver biopsy were performed. Results The prevalence of clinically significant liver fibrosis (F ≥ 2) was 35.1%. Alanine aminotransferase, aspartate aminotransferase and gamma-glutamyltransferase values were higher in the F ≥ 2 group. Metabolic syndrome was associated with the presence of clinically significant fibrosis (OR 14.2; 95% CI 3.7–55; p Conclusions Over one-third of asymptomatic and lung affected adults with ‘PI*ZZ’ AATD have significant underlying liver fibrosis. Liver biopsies demonstrated variable amounts of accumulated Z AAT. The risk of liver fibrosis increases in the presence of metabolic syndrome, accumulation of AAT in hepatocytes, and portal inflammation on baseline biopsy. The results support the hypothesis that liver disease in this genetic condition may be related to a “toxic gain of function” from accumulation of AAT in hepatocytes. Lay summary Individuals diagnosed with classic alpha-1 antitrypsin deficiency (ZZ) are at risk of liver injury and scarring, because of the accumulation of abnormal alpha-1 antitrypsin in the liver. A liver biopsy in ZZ individuals can demonstrate the accumulation of alpha-1 antitrypsin within the liver and identify if any associated liver scarring is present. Indviduals with large amounts of alpha-1 antitrypsin on biopsy may be at risk of liver injury and fibrosis. Additional common medical conditions of diabetes, obesity, high cholesterol, and hypertension (known as metabolic syndrome) are associated with a greater degree of liver injury. Clinical Trial number clinicaltrials.gov NCT01810458.

Despite the reduced caloric content of artificially sweetened beverages (ASBs) relative to those sweetened with sucrose, consumption of ASBs fail to consistently decrease the risk of obesity and associated diseases. This failure may be due to the inability of ASBs to effectively reduce appetite and hence overall caloric intake. A variety of non-nutritive sweeteners (NNS), however, remain to be screened for effectiveness in promoting satiety and reducing calorie consumption. Erythritol is well-tolerated, low-calorie sugar alcohol widely used as a sugar substitute. It is unique among NNS due to its low sweetness index relative to glucose, meaning that it is typically served at much higher concentrations than other common NNS. Animal and human studies have noted correlations between osmolarity, satiety, and levels of satiety hormones, independent of the effects of sweetness or nutritive value. We hypothesized that consumption of a beverage sweetened with erythritol to the sweetness and osmolarity of a common soft drink will improve self-reported satiety and more strongly affect the magnitude of changes in the hormone ghrelin than would an iso sweet beverage sweetened only with aspartame, a sweetener with a high sweetness index relative to glucose. Using a randomized double-blind crossover trial, we found that serum ghrelin was significantly decreased after consumption of an erythritol-sweetened beverage compared to aspartame. Likewise, consumption of the erythritol-sweetened beverage increased various measures of satiety in volunteers. Knowledge gained from this project demonstrates that high-osmolarity NNS may be useful in formulating ASBs that are satiating and low in calories.

Background Alpha-1 antitrypsin deficiency (AATD)-mediated liver disease is a toxic “gain-of-function” inflammation in the liver associated with intracellular retention of mutant alpha-1 antitrypsin. The clinical presentation of the disease includes fibrosis, cirrhosis and liver failure. However, the pathogenic mechanism of AATD-mediated liver disease is not well understood. Here, we investigated the role of plasma extracellular vesicles (EVs) in progression of AATD-mediated liver disease. Methods EVs were isolated from plasma of AATD individuals with liver disease and healthy controls. Their cytokines and miRNA content were examined by multiplex assay and small RNA sequencing. The bioactivity of EVs was assessed by qPCR, western blot analysis and immunofluorescent experiments using human hepatic stellate cells (HSCs) treated with EVs isolated from control or AATD plasma samples. Results We have found that AATD individuals have a distinct population of EVs with pathological cytokine and miRNA contents. When HSCs were cultured with AATD plasma derived-EVs, the expression of genes related to the development of fibrosis were significantly amplified compared to those treated with healthy control plasma EVs. Conclusion AATD individuals have a distinct population of EVs with abnormal cytokine and miRNA contents and the capacity to activate HSCs and mediate fibrosis. Better understanding of the components which cause liver inflammation and fibrogenesis, leading to further liver injury, has the potential to lead to the development of new treatments or preventive strategies to prevent AATD-mediated liver disease.

BACKGROUND Obesity has been associated with attenuated vaccine responses and an increased risk of contracting pneumococcal pneumonia, but no study to our knowledge has assessed the impact of obesity and genetics on 23-valent pneumococcal vaccine (PPSV23) efficacy. We assessed the relationship of obesity (primary analysis) and stimulator of interferon genes (STING1) genotype (secondary analysis) on PPSV23 efficacy. METHODS Nonobese (BMI 22–25 kg/m2) and obese participants (BMI ≥30 kg/m2) were given a single dose of PPSV23. Blood was drawn immediately prior to and 4–6 weeks after vaccination. Serum samples were used to assess PPSV23-specific antibodies. STING1 genotypes were identified using PCR on DNA extracted from peripheral blood samples. RESULTS Forty-six participants were categorized as nonobese (n = 23; 56.5% women; mean BMI 23.3 kg/m2) or obese (n = 23; 65.2% women; mean BMI 36.3 kg/m2). Obese participants had an elevated fold change in vaccine-specific responses compared with nonobese participants (P < 0.0001). The WT STING1 group (R232/R232) had a significantly higher PPSV23 response than individuals with a single copy of HAQ-STING1 regardless of BMI (P = 0.0025). When WT was assessed alone, obese participants had a higher fold serotype-specific response compared with nonobese participants (P < 0.0001), but no difference was observed between obese and nonobese individuals with 1 HAQ allele (P = 0.693). CONCLUSIONS These observations demonstrate a positive association between obesity and PPSV23 efficacy specifically in participants with the WT STING1 genotype. TRIAL REGISTRATION ClinicalTrials.gov NCT02471014. FUNDING This research was supported by the NIH and the University of Florida MD-PhD Training Program., Obesity and a WT STING1 genotype are positively associated with efficacy of the 23-valent pneumococcal vaccine in a small cohort of subjects.

The α-1-antitrypsin (or alpha-1-antitrypsin, A1AT) Z variant is the primary cause of severe A1AT deficiency and forms polymeric chains that aggregate in the endoplasmic reticulum of hepatocytes. Around 2%–5% of Europeans are heterozygous for the Z and WT M allele, and there is evidence of increased risk of liver disease when compared with MM A1AT individuals. We have shown that Z and M A1AT can copolymerize in cell models, but there has been no direct observation of heteropolymer formation in vivo. To this end, we developed a monoclonal antibody (mAb2H2) that specifically binds to M in preference to Z A1AT, localized its epitope using crystallography to a region perturbed by the Z (Glu342Lys) substitution, and used Fab fragments to label polymers isolated from an MZ heterozygote liver explant. Glu342 is critical to the affinity of mAb2H2, since it also recognized the mild S-deficiency variant (Glu264Val) present in circulating polymers from SZ heterozygotes. Negative-stain electron microscopy of the Fab2H2-labeled liver polymers revealed that M comprises around 6% of the polymer subunits in the MZ liver sample. These data demonstrate that Z A1AT can form heteropolymers with polymerization-inert variants in vivo with implications for liver disease in heterozygous individuals., A monoclonal antibody developed as a single-molecule probe has revealed pathological aggregates of α-1-antitrypsin capture the WT variant.

Alpha 1-antitrypsin deficiency (AATD) is the most common genetic cause of liver disease in children and is associated with early-onset chronic liver disease in adults. AATD associated liver injury is caused by hepatotoxic retention of polymerized mutant alpha 1-antitrypsin molecules within the endoplasmic reticulum. Currently, there is no curative therapy for AATD. In this study, we selected small molecules with the potential to bind mutant alpha 1-antitrypsin (Z-variant) to inhibit its accumulation in hepatocytes. We used molecular docking to select candidate compounds that were validated in cell and animal models of disease. A crystal structure of polymerized alpha 1-antitrypsin molecule was used as the basis for docking 139,735 compounds. Effects of the top scoring compounds were investigated in a cell model that stably expresses Z-variant alpha 1-antitrypsin and in PiZ mice expressing Z-variant human alpha 1-antitrypsin (Z-hAAT), encoded by SERPINA1*E342K. 4&prime, &prime, 5-(Methylenedioxy)-2-nitrocinnamic acid was predicted to bind cleaved alpha 1-antitrypsin at the polymerization interface, and observed to co-localize with Z-hAAT, increase Z-hAAT degradation, inhibit intracellular accumulation of Z-hAAT, and alleviate liver fibrosis.

Pulmonary hypertension (PH) complicates the care of patients with chronic lung disease, such as idiopathic pulmonary fibrosis (IPF), resulting in a significant increase in morbidity and mortality. Disease pathogenesis is orchestrated by unidentified myeloid-derived cells. We used murine models of PH and pulmonary fibrosis to study the role of circulating myeloid cells in disease pathogenesis and prevention. We administered clodronate liposomes to bleomycin-treated wild-type mice to induce pulmonary fibrosis and PH with a resulting increase in circulating bone marrow–derived cells. We discovered that a population of C-X-C motif chemokine receptor (CXCR) 2+ myeloid-derived suppressor cells (MDSCs), granulocytic subset (G-MDSC), is associated with severe PH in mice. Pulmonary pressures worsened despite improvement in bleomycin-induced pulmonary fibrosis. PH was attenuated by CXCR2 inhibition, with antagonist SB 225002, through decreasing G-MDSC recruitment to the lung. Molecular and cellular analysis of clinical patient samples confirmed a role for elevated MDSCs in IPF and IPF with PH. These data show that MDSCs play a key role in PH pathogenesis and that G-MDSC trafficking to the lung, through chemokine receptor CXCR2, increases development of PH in multiple murine models. Furthermore, we demonstrate pathology similar to the preclinical models in IPF with lung and blood samples from patients with PH, suggesting a potential role for CXCR2 inhibitor use in this patient population. These findings are significant, as there are currently no approved disease-specific therapies for patients with PH complicating IPF.

Background Pathological mutations in Alpha-1 Antitrypsin (AAT) protein cause retention of toxic polymers in the hepatocyte endoplasmic reticulum. The risk for cirrhosis in AAT deficiency is likely directly related to retention of these polymers within the liver. Polymers are classically identified on liver biopsy as inclusion bodies by periodic acid schiff staining after diastase treatment and immunohistochemistry. However, characterization of the polymer burden within a biopsy sample is limited to a semi-quantitative scale as described by a pathologist. Better methods to quantify polymer are needed to advance our understanding of pathogenesis of disease. Therefore, we developed a method to quantify polymer aggregation from standard histologic specimens. In addition, we sought to understand the relationship of polymer burden and other histologic findings to the presence of liver fibrosis. Methods Liver samples from a well-categorized AATD cohort were used to develop histo-morphometric tools to measure protein aggregation. Results Whole-slide morphometry reliably quantifies aggregates in AATD individuals. Despite very low levels of inclusions present (0–0.41%), accumulation of globules is not linear and is associated with higher fibrosis stages. Immunohistochemistry demonstrates that fibrosis is associated with polymer accumulation and not total AAT. A proportion of patients were found to be “heavy accumulators” with a polymer burden above the upper 25% of normal distribution. Males had significantly more liver inclusions and polymer than females. These measurements also highlight interrelated phenotypes of hepatocellular degeneration and autophagy in AATD liver disease. Conclusion Quantitative inclusion analysis measures AAT accumulation in liver biopsy specimens. Quantification of polymer may identify individuals at risk for progressive disease and candidates for therapeutic interventions. Furthermore, these methods may be useful for evaluating efficacy of drugs targeting accumulation of AAT.

This study evaluated the bioequivalence, safety, and immunogenicity of a new liquid formulation of human plasma–derived alpha1-proteinase inhibitor, Liquid Alpha1-PI, compared with the Lyophilized ...

Many individuals with Alpha-1 Antitrypsin Deficiency (AATD) are unaware of their diagnosis. In the absence of an AATD diagnosis, irreversible damage continues, and incorrect care is provided. Research demonstrates low levels of knowledge about AATD among health care providers. To address this ongoing issue, a short educational video was developed for health care providers with the goal of increasing knowledge and testing for AATD. A five-question test on the video material was developed. Invitations to participate in the study were sent via email to providers at both public teaching hospitals and private practices across the country. Respondents completed three parts online: pre-test, video, and post-test. To confirm retention of knowledge gained, providers who completed all three were invited to take the same test 3–6 months later. There were 683 providers who responded, and 213 completed all three portions; 105 of those providers completed the 3–6-months of follow-up testing. The average pre-tes...

α(1)-Antitrypsin deficiency (AATD) is an inherited disease characterized by emphysema and liver disease. AATD is most often caused by a single amino acid substitution at position 342 in the mature protein, resulting in the Z mutation of the AAT gene (ZAAT). This substitution is associated with misfolding and accumulation of ZAAT in the endoplasmic reticulum (ER) of hepatocytes, causing a toxic gain of function. ERdj3 is an ER luminal DnaJ homologue, which, along with calreticulin, directly interacts with misfolded ZAAT. We hypothesize that depletion of each of these chaperones will change the fate of ZAAT polymers. Our study demonstrates that calreticulin modulation reveals a novel ZAAT degradation mechanism mediated by exosomes. Using human PiZZ hepatocytes and K42, a mouse calreticulin-deficient fibroblast cell line, our results show ERdj3 and calreticulin directly interact with ZAAT in PiZZ hepatocytes. Silencing calreticulin induces calcium independent ZAAT–ERdj3 secretion through the exosome pathway. This co-secretion decreases ZAAT aggregates within the ER of hepatocytes. We demonstrate that calreticulin has an inhibitory effect on exosome-mediated ZAAT–ERdj3 secretion. This is a novel ZAAT degradation process that involves a DnaJ homologue chaperone bound to ZAAT. In this context, calreticulin modulation may eliminate the toxic gain of function associated with aggregation of ZAAT in lung and liver, thus providing a potential new therapeutic approach to the treatment of AATD-related liver disease.

There are a number of diseases characterized by the presence of neutrophil elastase (NE) activity in tissues including cystic fibrosis and alpha-1-antitrypsin deficiency induced lung destruction. It is generally accepted that NE actively contributes to this pathological process, but the precise mechanisms has yet to be determined. We hypothesized that NE activates the macrophages (M□) pro-inflammatory program. We demonstrate that following NE exposure, monocyte-derived M□ release proteolytic activity composed of several matrix metalloproteinases (MMPs) which could contribute to extracellular matrix (ECM) degradation. NE upregulates expression of M□ derived pro-inflammatory cytokines including TNFα, IL-1β, and IL-8. Thus, NE-activated M□ can contribute to tissue destruction through the proteolytic activity of metalloproteinases and by supporting chronic inflammation through expression of pro-inflammatory cytokines. We also demonstrate that NE increases M□ adhesion that is attenuated by antibodies specific to integrin subunits. We show that the effects of NE on M□ can be mediated through an activation of integrin pathways. In support of integrin involvement, we demonstrate that NE activates the Src kinase family, a hallmark of integrin signaling activation. Moreover, pretreatment of macrophages with a specific Src kinase inhibitor, PP2, completely prevents NE-induced inflammatory cytokine production. Taken together these findings indicate that NE has effect on lung destruction that extends beyond direct proteolytic degradation of matrix proteins.

Alpha-1 antitrypsin deficiency (AATD) is a largely monogenetic disorder associated with a high risk for the development of chronic obstructive pulmonary disease (COPD) and cirrhosis. Intravenous alpha-1 antitrypsin (AAT) therapy has been available for the treatment of individuals with AATD and COPD since the late 1980s. Initial Food and Drug Administration (FDA) approval was granted based on biochemical efficacy. Following its approval, the FDA, scientific community and third-party payers encouraged manufacturers of AAT therapy to determine its clinical efficacy. This task has proved challenging because AATD is a rare, orphan disorder comprised of individuals who are geographically dispersed and infrequently identified. In addition, robust clinical trial outcomes have been lacking until recently. This review provides an update on the evidence for the clinical efficacy of intravenous AAT therapy for patients with AATD-related emphysema.

Background: The diagnosis and clinical management of adults with alpha-1 antitrypsin deficiency (AATD) have been the subject of ongoing debate, ever since the publication of the first American Thoracic Society guideline statement in 1989.1 In 2003, the "American Thoracic Society (ATS)/European Respiratory Society (ERS) Statement: Standards for the Diagnosis and Management of Individuals with Alpha-1 Antitrypsin Deficiency" made a series of evidence-based recommendations, including a strong recommendation for broad-based diagnostic testing of all symptomatic adults with chronic obstructive pulmonary disease (COPD).2 Even so, AATD remains widely under-recognized. To update the 2003 systematic review and clinical guidance, the Alpha-1 Foundation sponsored a committee of experts to examine all relevant, recent literature in order to provide concise recommendations for the diagnosis and management of individuals with AATD. Purpose: To provide recommendations for: (1) the performance and interpretation of diagnostic testing for AATD, and (2) the current management of adults with AATD and its associated medical conditions. Methods: A systematic review addressing the most pressing questions asked by clinicians (clinician-centric) was performed to identify citations related to AATD that were published since the 2003 comprehensive review, specifically evaluating publications between January 2002 and December 2014. Important, more recent publications were solicited from the writing committee members as well. The combined comprehensive literature reviews of the 2003 document and this current review comprise the evidence upon which the committee's conclusions and recommendations are based. Results: Recommendations for the diagnosis and management of AATD were formulated by the committee. Conclusions: The major recommendations continue to endorse and reinforce the importance of testing for AATD in all adults with symptomatic fixed airflow obstruction, whether clinically labeled as COPD or asthma. Individuals with unexplained bronchiectasis or liver disease also should be tested. Family testing of first-degree relatives is currently the most efficient detection technique. In general, individuals with AATD and emphysema, bronchiectasis, and/or liver disease should be managed according to usual guidelines for these clinical conditions. In countries where intravenous augmentation therapy with purified pooled human plasma-derived alpha-1 antitrypsin is available, recent evidence now provides strong support for its use in appropriate individuals with lung disease due to AATD.

In Figure 6 of this article (PDF and HTML) the arrows labelled 'Depolymerization' and 'Polymerization' should be labelled 'Polymerization' and 'Depolymerization', respectively.

Alpha-1 antitrypsin deficiency (AATD) liver disease is characterized by marked heterogeneity in presentation and progression, despite a common underlying gene mutation, strongly suggesting the involvement of other genetic and/or epigenetic modifiers. Variation in clinical phenotype has added to the challenge of detection, diagnosis, and testing of new therapies in patients with AATD. We examined the contribution of DNA methylation (5-methylcytosine [5mC]) to AATD liver disease heterogeneity because 5mC responds to environmental and genetic cues and its deregulation is a major driver of liver disease. Using liver biopsies from adults with early-stage AATD and the ZZ genotype, genome-wide 5mC patterns were interrogated. We compared DNA methylation among patients with early AATD, and among patients with normal liver, cirrhosis, and hepatocellular carcinoma derived from multiple etiologic exposures, and linked patient clinical/demographic features. Global analysis revealed significant genomic hypomethylation in AATD liver-impacting genes related to liver cancer, cell cycle, and fibrosis, as well as key regulatory molecules influencing growth, migration, and immune function. Further analysis indicated that 5mC changes are localized, with hypermethylation occurring within a background of genome-wide 5mC loss and with patients with AATD manifesting distinct epigenetic landscapes despite their mutational homogeneity. By integrating clinical data with 5mC landscapes, we observed that CpGs differentially methylated among patients with AATD disease are linked to hallmark clinical features of AATD (e.g., hepatocyte degeneration and polymer accumulation) and further reveal links to well-known sex-specific effects of liver disease progression. Conclusion: Our data reveal molecular epigenetic signatures within this mutationally homogeneous group that point to ways to stratify patients for liver disease risk.