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Introduction: A previous 2-week clinical trial of aerosolized hyaluronan (HA) in COPD showed a rapid reduction in lung elastic fiber breakdown, as measured by sputum levels of the unique elastin crosslinks, desmosine and isodesmosine (DID). To further assess the therapeutic efficacy of HA and the utility of DID as surrogate markers for the development of pulmonary emphysema, we have conducted a 28-day randomized, double-blind, placebo-controlled, phase 2 trial of HA involving 27 subjects with alpha-1 antiprotease deficiency COPD., Methods: The study drug consisted of a 3 ml inhalation solution containing 0.03% HA with an average molecular weight of 150 kDa that was self-administered twice daily. DID levels were measured in urine, sputum, and plasma using tandem mass spectrometry., Results: Free urine DID in the HA group showed a significant negative correlation with time between days 14 and 35 (r = -1.0, p = 0.023) and was statistically significantly decreased from baseline at day 35 (15.4 vs 14.2 ng/mg creatinine, p = 0.035). A marked decrease in sputum DID was also seen in the HA group between days 1 and 28 (0.96 vs 0.18 ng/mg protein), but the difference was not significant, possibly due to the small number of adequate specimens. Plasma DID remained unchanged following HA treatment and no significant reductions in urine, sputum, or plasma DID were seen in the placebo group., Conclusions: The results support additional clinical trials to further evaluate the therapeutic effect of HA and the use of DID as a real-time marker of drug efficacy., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Introduction: The Multicenter International Lymphangioleiomyomatosis (LAM) Efficacy of Sirolimus (MILES) trial revealed that sirolimus stabilised lung function in patients with moderately severe LAM. The purpose of this study was to further examine the MILES cohort for the effects of racial, demographic, clinical and physiological patient characteristics on disease progression and treatment response in LAM., Methods: MILES subjects were stratified on the basis of menopausal status (pre-menopausal/post-menopausal), race (Asian/Caucasian), bronchodilator responsiveness (present/absent), initial forced expiratory volume in 1 s (FEV 1 ; 51-70% versus ≤50% predicted) and tuberous sclerosis complex (TSC) association (yes/no). A linear mixed effects model was used to compare slope differences, and nonparametric tests were used to compare medians and proportions between treatment groups in each stratum., Results: In the MILES placebo group, pre-menopausal patients declined 5-fold faster than post-menopausal patients (mean±se FEV 1 slope -17±3 versus -3±3 mL·month -1 ; p=0.003). Upon treatment with sirolimus, both the pre-menopausal (-17±3 versus -1±2 mL·month -1 ; p<0.0001) and post-menopausal patients (-3±3 versus 6±3 mL·month -1 ; p=0.04) exhibited a beneficial response in mean±se FEV 1 slope compared with the placebo group. Race, LAM subtype, bronchodilator responsiveness or baseline FEV 1 did not impact the rate of disease progression in the placebo group or treatment response in the sirolimus group. Menopausal status and race had differential effects on the adverse event profile of sirolimus. Baseline serum vascular endothelial growth factor (VEGF)-D >600 pg·mL -1 identified subgroups of patients who were more likely to decline on placebo and respond to treatment with sirolimus., Conclusions: In LAM patients, treatment with sirolimus is beneficial regardless of menopausal status, race, bronchodilator responsiveness, baseline FEV 1 or TSC association. Serum VEGF-D and menopausal status can help inform therapeutic decisions., Competing Interests: Conflict of interest: N. Gupta has nothing to disclose. Conflict of interest: H-S. Lee has nothing to disclose. Conflict of interest: L.R. Young reports advisory board work for Boehringer Ingelheim and royalties for authorship from UpToDate, outside the submitted work; and has a patent Serum VEGF-D, no royalties issued. Conflict of interest: C. Strange reports grants for studies of LAM from Novartis, outside the submitted work. Conflict of interest: J. Moss has nothing to disclose. Conflict of interest: L.G. Singer has nothing to disclose. Conflict of interest: K. Nakata has nothing to disclose. Conflict of interest: A.F. Barker has nothing to disclose. Conflict of interest: J.T. Chapman has nothing to disclose. Conflict of interest: M.L. Brantly has nothing to disclose. Conflict of interest: J.M. Stocks has nothing to disclose. Conflict of interest: K.K. Brown reports grants from NHLBI, personal fees from AstraZeneca, Biogen, Galecto, MedImmune, Novartis, ProMetic, Patara, Third Pole, Galapagos, Boehringer Ingelheim, Theravance and Three Lakes Partners, conversation under CDA only from Genoa, other (submitted grant) from Roche/Genentech, outside the submitted work. Conflict of interest: J.P. Lynch has nothing to disclose. Conflict of interest: H.J. Goldberg has nothing to disclose. Conflict of interest: G.P. Downey has nothing to disclose. Conflict of interest: A.M. Taveira-DaSilva has nothing to disclose. Conflict of interest: J.P. Krischer has nothing to disclose. Conflict of interest: K. Setchell has nothing to disclose. Conflict of interest: B.C. Trapnell has nothing to disclose. Conflict of interest: Y. Inoue reports grants from Japanese Ministry of Health, Labor, and Welfare, during the conduct of the study. Conflict of interest: F.X. McCormack has a patent on serum VEGF-D testing. All royalties are waived to the parent institution, the University of Cincinnati., (The content of this work is not subject to copyright. Design and branding are copyright ©ERS 2019.)

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., Competing Interests: The work of this writing committee was supported by the Alpha-1 Foundation through payment of travel and meeting expenses and payment of library searching and printing fees. No other assistance was provided. Specifically, there was not honoraria paid and there was not writing assistance of any sort provided. Ms. Knight and Drs. Turino, Brantly, and Goodman have nothing to disclose. Dr. Sandhaus reports grants from CSL Behring. He is employed part-time by the Alpha-1 Foundation and AlphaNet, 2 not-for-profit entities that support research and health management in alpha-1 antitrypsin deficiency and receive part of their funding from pharmaceutical industry sources. Dr. Sandhaus serves on the Medical and Scientific Advisory Committee of the COPD Foundation. Dr. Campos reports grants from Grifols and CSL Behring. Dr. Cross reports personal fees from CSL Behring and Baxter. Dr. Hogarth reports personal fees from Baxter, CSL Behring, and Grifols. Dr. Stocks reports grants to the University of Texas from Kamada, Grifols, and Talecris. Dr. Stoller has served as a scientific consultant to Grifols, CSL Behring, Baxalta, and Kamada regarding alpha-1 antitrypsin deficiency and serves on the Board of Directors of the Alpha-1 Foundation and on the Medical and Scientific Advisory Committees of the Alpha-1 Foundation and the COPD Foundation. Dr. Strange reports personal fees and stock options from Abeona, grants from Baxalta, grants and personal fees from CSL Behring, grants and personal fees from Grifols, and grants from Alpha-1 Foundation. Dr. Teckman reports he is a paid consultant for Isis Pharmaceuticals, Arrowhead Research, Alnylam Inc., Editas Inc., Proteostasis Inc., Genkyotix, Grifols, Intellia, Retrophin, RxCelerate, Velfene, and Vertex. He has received grants from the National Institutes of Health and the Alpha-1 Foundation.

Alpha-1 antitrypsin (AAT) deficiency and tobacco smoking are confirmed risk factors for Chronic Obstructive Pulmonary Disease. We hypothesized that variable DNA methylation would be associated with smoking and inflammation, as reflected by the level of C-Reactive Protein (CRP) in AAT-deficient subjects. Methylation levels of 1,411 autosomal CpG sites from the Illumina GoldenGate Methylation Cancer Panel I were analyzed in 316 subjects. Associations of five smoking behaviors and CRP levels with individual CpG sites and average methylation levels were assessed using non-parametric testing, linear regression and linear mixed effect models, with and without adjustment for age and gender. Univariate linear regression analysis revealed that methylation levels of 16 CpG sites significantly associated with ever-smoking status. A CpG site in the TGFBI gene was the only site associated with ever-smoking after adjustment for age and gender. No highly significant associations existed between age at smoking initiation, pack-years smoked, duration of smoking, and time since quitting smoking as predictors of individual CpG site methylation levels. However, ever-smoking and younger age at smoking initiation associated with lower methylation level averaged across all sites. DNA methylation at CpG sites in the RUNX3, JAK3 and KRT1 genes associated with CRP levels. The most significantly associated CpG sites with gender and age mapped to the CASP6 and FZD9 genes, respectively. In summary, this study identified multiple potential candidate CpG sites associated with ever-smoking and CRP level in AAT-deficient subjects. Phenotypic variability in Mendelian diseases may be due to epigenetic factors.

Background: The development of COPD in subjects with alpha-1 antitrypsin (AAT) deficiency is likely to be influenced by modifier genes. Genome-wide association studies and integrative genomics approaches in COPD have demonstrated significant associations with SNPs in the chromosome 15q region that includes CHRNA3 (cholinergic nicotine receptor alpha3) and IREB2 (iron regulatory binding protein 2).We investigated whether SNPs in the chromosome 15q region would be modifiers for lung function and COPD in AAT deficiency., Methods: The current analysis included 378 PIZZ subjects in the AAT Genetic Modifiers Study and a replication cohort of 458 subjects from the UK AAT Deficiency National Registry. Nine SNPs in LOC123688, CHRNA3 and IREB2 were selected for genotyping. FEV1 percent of predicted and FEV1/FVC ratio were analyzed as quantitative phenotypes. Family-based association analysis was performed in the AAT Genetic Modifiers Study. In the replication set, general linear models were used for quantitative phenotypes and logistic regression models were used for the presence/absence of emphysema or COPD., Results: Three SNPs (rs2568494 in IREB2, rs8034191 in LOC123688, and rs1051730 in CHRNA3) were associated with pre-bronchodilator FEV1 percent of predicted in the AAT Genetic Modifiers Study. Two SNPs (rs2568494 and rs1051730) were associated with the post-bronchodilator FEV1 percent of predicted and pre-bronchodilator FEV1/FVC ratio; SNP-by-gender interactions were observed. In the UK National Registry dataset, rs2568494 was significantly associated with emphysema in the male subgroup; significant SNP-by-smoking interactions were observed., Conclusions: IREB2 and CHRNA3 are potential genetic modifiers of COPD phenotypes in individuals with severe AAT deficiency and may be sex-specific in their impact.

Alpha-1-antitrypsin deficiency is a genetic condition associated with severe, early-onset chronic obstructive pulmonary disease (COPD). However, there is significant variability in lung function impairment among persons with the protease inhibitor ZZ genotype. Early identification of persons at highest risk of developing lung disease could be beneficial in guiding monitoring and treatment decisions. Using a multicenter, family-based study sample (2002-2005) of 372 persons with the protease inhibitor ZZ genotype, the authors developed prediction models for forced expiratory volume in 1 second (FEV(1)) and the presence of severe COPD using demographic, clinical, and genetic variables. Half of the data sample was used for model development, and the other half was used for model validation. In the training sample, variables found to be predictive of both FEV(1) and severe COPD were age, sex, pack-years of smoking, bronchodilator responsiveness, chronic bronchitis symptoms, and index case status. In the validation sample, the predictive model for FEV(1) explained 50% of the variance in FEV(1), and the model for severe COPD exhibited excellent discrimination (c statistic = 0.88).

Severe alpha(1)-antitrypsin (AAT) deficiency is a proven genetic risk factor for chronic obstructive pulmonary disease (COPD), especially in individuals who smoke. There is marked variability in the development of lung disease in individuals homozygous (PI ZZ) for this autosomal recessive condition, suggesting that modifier genes could be important. We hypothesized that genetic determinants of obstructive lung disease may be modifiers of airflow obstruction in individuals with severe AAT deficiency. To identify modifier genes, we performed family-based association analyses for 10 genes previously associated with asthma and/or COPD, including IL10, TNF, GSTP1, NOS1, NOS3, SERPINA3, SERPINE2, SFTPB, TGFB1, and EPHX1. All analyses were performed in a cohort of 378 PI ZZ individuals from 167 families. Quantitative spirometric phenotypes included forced expiratory volume in one second (FEV(1)) and the ratio of FEV(1)/forced vital capacity (FVC). A qualitative phenotype of moderate-to-severe COPD was defined for individuals with FEV(1)

Background: Severe alpha(1)-antitrypsin (AAT) deficiency is an autosomal recessive genetic condition associated with an increased but variable risk for chronic obstructive pulmonary disease (COPD). A study was undertaken to assess the impact of chronic bronchitis, pneumonia, asthma and sex on the development of COPD in individuals with severe AAT deficiency., Methods: The AAT Genetic Modifier Study is a multicentre family-based cohort study designed to study the genetic and epidemiological determinants of COPD in AAT deficiency. 378 individuals (age range 33-80 years), confirmed to be homozygous for the SERPINA1 Z mutation, were included in the analyses. The primary outcomes of interest were a quantitative outcome, forced expiratory volume in 1 s (FEV(1)) percentage predicted, and a qualitative outcome, severe airflow obstruction (FEV(1) <50% predicted)., Results: In multivariate analysis of the overall cohort, cigarette smoking, sex, asthma, chronic bronchitis and pneumonia were risk factors for reduced FEV(1 )percentage predicted and severe airflow obstruction (p<0.01). Index cases had lower FEV(1) values, higher smoking histories and more reports of adult asthma, pneumonia and asthma before age 16 than non-index cases (p<0.01). Men had lower pre- and post-bronchodilator FEV(1) percentage predicted than women (p<0.0001); the lowest FEV(1) values were observed in men reporting a history of childhood asthma (26.9%). This trend for more severe obstruction in men remained when index and non-index groups were examined separately, with men representing the majority of non-index individuals with airflow obstruction (71%). Chronic bronchitis (OR 3.8, CI 1.8 to 12.0) and a physician's report of asthma (OR 4.2, CI 1.4 to 13.1) were predictors of severe airflow obstruction in multivariate analysis of non-index men but not women., Conclusion: In individuals with severe AAT deficiency, sex, asthma, chronic bronchitis and pneumonia are risk factors for severe COPD, in addition to cigarette smoking. These results suggest that, in subjects severely deficient in AAT, men, individuals with symptoms of chronic bronchitis and/or a past diagnosis of asthma or pneumonia may benefit from closer monitoring and potentially earlier treatment.

We measured intra- and extravascular body-fluid compartments in 12 resting males before (day 1; control), during (day 8) and after (day 22) a 3-week, exercise-heat acclimation protocol to investigate plasma volume (PV) changes. Our specific focus was upon the selective nature of the acclimation-induced PV expansion, and the possibility that this expansion could be sustained during prolonged acclimation. Acclimation was induced by cycling in the heat, and involved 16 treatment days (controlled hyperthermia (90 min); core temperature = 38.5 degrees C) and three experimental exposures (40 min rest, 96.9 min (s.d. 9.5 min) cycling), each preceded by a rest day. The environmental conditions were a temperature of 39.8 degrees C (s.d. 0.5 degrees C) and relative humidity of 59.2% (s.d. 0.8%). On days 8 and 22, PV was expanded and maintained relative to control values (day 1: 44.0 +/- 1.8; day 8: 48.8 +/- 1.7; day 22: 48.8 +/- 2.0 ml kg(-1); P < 0.05). The extracellular fluid compartment (ECF) was equivalently expanded from control values on days 8 (279.6 +/- 14.2 versus 318.6 +/- 14.3 ml kg(-1); n= 8; P < 0.05) and 22 (287.5 +/- 10.6 versus 308.4 +/- 14.8 ml kg(-1); n= 12; P < 0.05). Plasma electrolyte, total protein and albumin concentrations were unaltered following heat acclimation (P > 0.05), although the total plasma content of these constituents was elevated (P < 0.05). The PV and interstitial fluid (ISF) compartments exhibited similar relative expansions on days 8 (15.0 +/- 2.2%versus 14.7 +/- 4.1%; P > 0.05) and 22 (14.4 +/- 3.6% versus 6.4 +/- 2.2%; P= 0.10). It is concluded that the acclimation-induced PV expansion can be maintained following prolonged heat acclimation. In addition, this PV expansion was not selective, but represented a ubiquitous expansion of the extracellular compartment.

We tested the hypothesis that local sweat rates would not display a systematic postadaptation redistribution toward the limbs after humid heat acclimation. Eleven nonadapted males were acclimated over 3 wk (16 exposures), cycling 90 min/day, 6 days/wk (40 degrees C, 60% relative humidity), using the controlled-hyperthermia acclimation technique, in which work rate was modified to achieve and maintain a target core temperature (38.5 degrees C). Local sudomotor adaptation (forehead, chest, scapula, forearm, thigh) and onset thresholds were studied during constant work intensity heat stress tests (39.8 degrees C, 59.2% relative humidity) conducted on days 1, 8, and 22 of acclimation. The mean body temperature (Tb) at which sweating commenced (threshold) was reduced on days 8 and 22 (P < 0.05), and these displacements paralleled the resting thermoneutral Tb shift, such that the Tb change to elicit sweating remained constant from days 1 to 22. Whole body sweat rate increased significantly from 0.87 +/- 0.06 l/h on day 1 to 1.09 +/- 0.08 and 1.16 +/- 0.11 l/h on days 8 and 22, respectively. However, not all skin regions exhibited equivalent relative sweat rate elevations from day 1 to day 22. The relative increase in forearm sweat rate (117 +/- 31%) exceeded that at the forehead (47 +/- 18%; P < 0.05) and thigh (42 +/- 16%; P < 0.05), while the chest sweat rate elevation (106 +/- 29%) also exceeded the thigh (P < 0.05). Two unique postacclimation observations arose from this project. First, reduced sweat thresholds appeared to be primarily related to a lower resting Tb, and more dependent on Tb change. Second, our data did not support the hypothesis of a generalized and preferential trunk-to-limb sweat redistribution after heat acclimation.

Objective: To assess the efficacy, safety of cefaclor for the treatment of pharyngotonsillitis in Pakistani children., Method: Pakistani children between the ages of two to twelve years and diagnosed with pharyngotonsillitis were eligible to participate in the study. Cefaclor was administered three times daily for a total dose of 20-40 mg/kg/day, depending on the severity of the infection, for 7 days. Patients were evaluated on the basis of symptomatic response and antibiotic susceptibility pattern as a result of bacteriological examination., Results: Based on observed pre-therapy signs/symptoms of infection with pharyngotonsillitis, 99% of patients (95% CI=94.9% to 100.0%) were reported as having a symptomatic response of Cure or Improvement by end of study. Group A b-haemolytic Streptococci was the most prevalent pathogen isolated pre-therapy (occurring in 45% of patients). Of all patients for whom this pathogen was isolated, 64% had complete eradication by end of therapy, while there was persistence or superinfection in 36% of patients. Only 9 occurrences of non-serious adverse events were observed among 7 of the 107 patients, including diarrhea (5), nausea (1), vomiting (1), rash (1) and stomatitis (1)., Conclusion: Based on the high symptomatic response rate of 99% and the positive safety data, cefaclor was found to be effective and safe in treating Pakistani children with pharyngotonsillitis.

This project examined the effects of repeated, resting cold-water immersion on metabolic heat production and core temperature defence during subsequent rest and exercising immersions. Seven males undertook 15 days of cold-water adaptation, immersed to the fourth intercostal space, with cold-water stress tests (CWST) on days 1, 8 and 15 (18.1 SD 0.1 degree C: 60 min seated, followed by 30 min cycling (1 W.kg-1)), and 90-min resting immersions (18.4 SD 0.4 degree C) on each of the intervening days. Adaptation elicited an habituated thermogenic response during the rest phase of CWST3 beyond 20 min, compared to CWST1 (P < 0.05), with oxygen consumption averaging 11.15 (+/- 0.25) ml.kg-1.min-1 and 8.61 (+/- 0.90) ml.kg-1.min-1 by 50 min, for CWST1 and CWST3, respectively. During exercise, this metabolic blunting was only apparent over the first 10-min period (60-70 min). No significant differences were observed during either the rest or exercise phases of the CWSTs for oesophageal temperature (Tes). While repeated cold-water exposures produced an habituated-thermogenic response, for an equivalent drop in Tes during rest, neither this response, nor an elevated thermogenesis, were apparent during subsequent cold-water exercise.

Classical ferruginous bodies in tissue samples are considered to be markers of past exposure to asbestos. Recent studies have shown that the presence of ferruginous bodies in bronchoalveolar lavage (BAL) fluid correlates with past exposure to asbestos and offers a more sensitive reference than occupational history. Lavage samples from five subjects who had worked in foundries were evaluated by light microscopy for the presence of ferruginous bodies and by transmission electron microscopy for both characterisation of the uncoated fibre burden and analysis of the cores of the ferruginous bodies. All samples at lower magnification (light microscopy (200 x)) contained ferruginous bodies that were externally consistent with asbestos bodies. At higher magnification (400 x), a separate population from this group could be identified by the presence of a thin black ribbon. Transmission electron microscopy of the core materials of ferruginous bodies and comparable uncoated particulates supported the reliability of higher magnification light microscopy for distinguishing most of those non-asbestos cores; however, a population of transparent non-asbestos cored ferruginous bodies were also shown to exist.

The high Mr glycoprotein (gp300) of equine herpesvirus type 1 was found to have an Mr, estimated by SDS-PAGE, of over 400,000 and was confirmed as being a surface glycoprotein by 125I-labelling. In contrast to [3H]glucosamine, gp300 showed very low levels of [3H]glucosamine, gp300 showed very low levels of [3H]mannose incorporation. The Mr of gp300 showed no detectable change upon treatment of purified virus with N-glycanase, and showed only a small change in virus-infected cells treated with tunicamycin. In addition, gp300 failed to bind the lectin concanavalin A. Taken together, these results indicate a lack of N-linked carbohydrate on gp300. The major carbohydrate species were found to be composed primarily of O-linked chains, as indicated by the sensitivity of the protein to monensin, to exoglycanase enzymes specific for sugars present in O-linked chains and to mild alkaline borohydride treatment, which revealed three species of carbohydrate of Mr of greater than 10,000, 2400 and 1100, respectively. Neuraminidase treatment and binding of Helix pomatia lectin indicated the presence of alpha-N-acetylglucosamine and sialic acid as terminal sugars. Immunological cross-reactivity of gp300 with a high Mr protein of equine herpesvirus type 4 was shown and it also exhibited a marked Mr variation in the vaccine strain Rhinomune.

Equine herpesvirus types 1 and 4 (EHV-1 and EHV-4) labelled with [14C]glucosamine were purified from infected cell culture medium and profiles of their structural proteins were obtained that enabled identification of the major glycoproteins. Nine glycosylated polypeptides were identified for each virus. Preparations of the purified viruses each contained a glycoprotein which was linked by disulphide bonds, as determined by diagonal gel electrophoresis under reducing/non-reducing conditions. High Mr forms of this glycoprotein were detected for EHV-1 when the sample was not heated. The EHV-1 protein consisted of three polypeptides of Mr 108K, 76K and 58K and the EHV-4 protein consisted of three polypeptides of Mr 112K, 74K and 61K. Western blotting and immunoprecipitation with monoclonal antibodies confirmed that the EHV-1 gB homologue migrates with an apparent Mr of 108K (140K under non-reducing conditions) but is cleaved to give glycoproteins of 76K and 58K which are held together by disulphide bonds. The EHV-4 gB homologue consists of a 112K glycoprotein which is cleaved to give glycoproteins of 74K and 61K which are also linked by disulphide bonds.