Study title: A systematic review of RCTs to examine the risk of adverse cardiovascular events with nicotine use.

Associations between cigarette smoking and increased risk of cardiovascular disease are well established. However, it is unclear whether the association is mediated by exposure to nicotine and/or to other constituents in cigarette smoke. The objective of this systematic review and meta-analysis of randomized control trials (RCTs) was to identify any potential associations between exposure to nicotine and the risk of clinically diagnosed adverse cardiovascular events in adult current users and nonusers of tobacco products. Among 1,996 results, 42 studies, comparing nicotine and non-nicotine groups, were included and were both qualitatively and quantitatively synthesized across the outcomes of arrhythmia, nonfatal myocardial infarction, nonfatal stroke, and cardiovascular death. The majority of studies evaluating nonfatal myocardial infarction, nonfatal stroke, and cardiovascular death reported no events that occurred in either the nicotine or non-nicotine control groups. Among the studies that reported events, rates of adverse events were similarly low between both groups. Consistent with findings from previous systematic reviews and meta-analyses, pooled data showed that rates for arrhythmia, nonfatal myocardial infarction, nonfatal stroke, and cardiovascular death were not significantly different between nicotine and non-nicotine groups. The overall quality of the body of evidence for each of the four outcomes of interest was graded as "moderate," limited only by the imprecision of results. The findings of this systematic review and meta-analysis indicate that, with moderate certainty, there are no significant associations between the use of nicotine and the risk of clinically diagnosed adverse cardiovascular events-specifically, arrhythmia, nonfatal myocardial infarction, nonfatal stroke, and cardiovascular death.

A systematic review of preclinical studies evaluating the association between nicotine and the initiation and progression of cancer.

Background: The association between cigarette smoking and the increased risk of many cancers is well established. Conversely, epidemiological studies of smokeless tobacco demonstrate decreased risk, or no elevated risk, of certain cancers versus smoking. However, it is unclear what role, if any, nicotine plays in these associations. The objective of this systematic review was to synthesize the available evidence from preclinical studies that examined the potential association between nicotine and the initiation and/or progression of cancer. Methods: MEDLINE, Embase, PsychInfo, and Cochrane Database of Systematic Reviews were searched for articles published from inception until February 13, 2022. Studies were eligible for inclusion if they evaluated animal cancer or tumor models, compared nicotine and non-nicotine groups, and evaluated measures of cancer initiation or progression. Results: Among 1,137 identified articles, 61 were included in qualitative synthesis. Twelve studies reported data on tumor initiation, and 54 studies reported data on tumor progression. The majority of the tumor initiation studies did not identify an association between nicotine exposure and an increased risk of spontaneous tumor initiation. Results of tumor progression studies were inconsistent and varied across the reported measures, cancer type being evaluated, and animal cancer model used. Overall, the quality of reporting was poor, with many studies not demonstrating a high level of internal and/or external validity. Conclusions: In conclusion, although animal models have provided invaluable data for human health risk assessments of chemical exposures, the heterogeneity across the studies included in this systematic review make the interpretation and generalizability of the results difficult.

Characterizing the gut microbiota in adults with bipolar disorder: a pilot study.

Background: Convergent evidence implicates gut microbiota in human health and disease. Hitherto, relatively few studies have evaluated the gut microbiota profile in individuals with bipolar disorder (BD) relative to healthy controls (HC). Methods: Fecal samples were collected from subjects (aged 18-65) meeting DSM-5-defined criteria for BD and age- and sex-matched HC without current or past history of mental or major medical disorders. Samples were sequenced using Illumina sequencing and association of specific taxa and co-occurrence of taxa with sample groups including the effect of diet was assessed using cluster analysis and analysis of communities of microorganisms (ANCOM). Nutritional composition was evaluated using the Dietary Questionnaire for Epidemiological Studies (DQES v2) Food Frequency Questionnaire. Results: Forty-six subjects were enrolled (n=23 BD, n=23 HC). Cluster analyses did not identify any significant differences between BD and HC (p=0.38). Lower microbiota diversity was observed among BD subjects relative to HC (p=0.04). A greater abundance of a Clostridiaceae OTU was observed among BD subjects when compared to HC and of Collinsella among BD-II subjects relative to BD-I. Cluster analysis revealed that neither diagnosis (p=0.38) nor diet (p=0.43) had a significant effect on overall gut microbiota composition. Limitations: This study has a small sample size and insufficient control for some potential moderating factors (e.g. psychotropic medication and smoking). Conclusion: This study suggests that individuals with BD may have a distinct gut microbiota profile compared to healthy controls, with a greater abundance of Clostridiaceae and Collinsella. These findings need to be replicated in future studies with larger sample sizes.

The murine female intestinal microbiota does not shift throughout the estrous cycle.

Pregnancy is accompanied by maternal physiological adaptations including metabolic, endocrine, immune, cardiovascular, skeletomuscular and neurological modifications that facilitate fetal and placental growth and development. Emerging evidence suggests that the maternal intestinal microbiota is modified over the course of healthy pregnancy. We have recently identified a maternal intestinal microbial shift within hours of conception; a shift that continued with advancing gestation. It is possible that maternal gut bacterial profiles might be associated with the known endocrine changes that accompany the female reproductive (estrous) cycle. METHODS: To determine whether the estrous cycle influenced the shifts in the maternal intestinal microbiota, time-matched fecal pellets were collected daily for 3 consecutive estrous cycles from individually housed, non-pregnant female C57BL/6J mice (n = 10) fed a control diet. Estrous stage was identified by cell type predominance in vaginal cytological samples. The corresponding fecal pellets for each estrous stage were processed for bacterial 16S rRNA sequencing of the variable 3 (V3) region. RESULTS: Estrous cycle stage accounted for a very small and not statistically significant proportion of the variation in the fecal microbiota according to PERMANOVA testing performed on Bray-Curtis dissimilarity scores. These values displayed no significant clustering of fecal microbial communities by estrous stage. CONCLUSION: The estrous cycle does not result in any significant shift in the intestinal microbial community in the reproductively mature, regularly cycling female mouse.

Erratum to: Do epinephrine auto-injectors have an unsuitable needle length in children and adolescents at risk for anaphylaxis from food allergy?

[This corrects the article DOI: 10.1186/s13223-016-0110-8.].

Fire Blight Resistance in Wild Accessions of Malus sieversii.

Fire blight (Erwinia amylovora) is a devastating bacterial disease in apple that results in severe economic losses. Epidemics are becoming more common as susceptible cultivars and rootstocks are being planted, and control is becoming more difficult as antibiotic-resistant strains develop. Resistant germplasm currently being utilized by breeding programs tend to have small fruit size and poor flavor characteristics. Malus sieversii, a progenitor species of domestic apple, is notable for its relatively large, palatable fruit and some accessions have been reported to be resistant to fire blight. In this study, nearly 200 accessions of M. sieversii and appropriate controls were inoculated with E. amylovora in both Washington and West Virginia to identify fire blight resistant accessions. Twelve accessions were identified with resistance comparable to highly resistant and resistant controls. Several accessions exhibited a unique resistance response, not previously reported in domestic apple (M. × domestica), characterized by low incidence of infection but high severity once infection was initiated. Several of these M. sieversii accessions will be used as parents in future crosses in the Washington State University apple breeding program.

Do epinephrine auto-injectors have an unsuitable needle length in children and adolescents at risk for anaphylaxis from food allergy?

BACKGROUND: Food allergy is the most common cause of anaphylaxis in children. Intramuscular delivery of epinephrine auto-injectors (EAI) is the standard of care for the treatment of anaphylaxis. We examined if children and adolescents at risk of anaphylaxis weighing 15-30 kg and >30 kg would receive epinephrine into the intramuscular space with the currently available EAI in North America and Europe. METHODS: The distance from skin to muscle (STMD) and skin to bone (STBD) on the mid third anterolateral area of the right thigh was measured by ultrasound applying either high pressure (max) or slight pressure (min) in 102 children weighing 15-30 kg (group 1) and 100 children and adolescents, weighing more than 30 kg (group 2). RESULTS: Using a high pressure EAI (HPEAI), Epipen Jr(®) and Auvi-Q(®)/Allerject(®) 0.15 mg, 11/102 (11 %) children in group 1 and 38/102 (38 %) using another HPEAI, Jext(®), had a STMDmax that showed a risk of intraosseous injection. There was a 1 % risk of subcutaneous injection with these devices. There was no risk of intraosseous injection using a low pressure EAI (LPEAI), Emerade(®). In group 2, the risk of intraosseous injection using a HPEAI was 3 % and no risk using a LPEAI. However, the risk of subcutaneous injection using HPEAI was 9 % and using LPEAI was 2 %. CONCLUSION: There is a risk of intraosseous injection using HPEAI (Epipen(®)/Epipen Jr(®), Auvi-Q(®)/Allerject(®) and especially Jext(®)) in children at risk of anaphylaxis. There was also a risk of subcutaneous injection using the currently available HPEAI in children and adolescents.

Toxicological evaluation of smokeless tobacco: 2-year chronic toxicity and carcinogenicity feeding study in Wistar Han rats.

UNLABELLED: A comprehensive 2-year oral chronic toxicity/carcinogenicity study was conducted with smokeless tobacco using modern toxicological test methods and well-accepted standards. The study included a 1-year interim subgroup to assess toxicity at that intermediate time point. Test groups consisted of a tobacco blend (B) used in snus, and an aqueous tobacco extract of that tobacco blend (E) administered at 0.2, 2, or 5 mg nicotine/kg body weight/day via dosed feed to male and female Wistar Han rats. The dosages were selected to simulate potential exposure in humans ingesting smokeless tobacco or an aqueous extract of smokeless tobacco (the latter intended to simulate a snus extract, to enable bridging these data to snus epidemiology data). The following endpoints were evaluated: clinical observations, body weights, feed consumption (FC), ophthalmic exams, toxicokinetics, clinical pathology, gross pathology, and histopathology. During the 2-year study, clear treatment-related, dose-responsive effects included: (1) increases in plasma nicotine and cotinine (indicating that animals were appropriately exposed to levels relevant to human exposure) and (2) decreases in body weights with some alterations in FC. At the 2-year time point, two tumor types (in the highest B doses) displayed statistically significantly increased incidence trends vs. CONTROLS: (1) uterine carcinoma in females and (2) epididymal mesothelioma in males. Three tumor types displayed statistically significantly decreased incidence trends: (1) mammary gland adenomas in females, (2) skin basal cell carcinomas in females, and (3) thyroid follicular cell adenomas in males. These increases (and decreases) in tumor trends were interpreted as not being treatment-related because: (1) there were no preneoplastic or related non-neoplastic histopathological findings in the treated rats at the 1-year or 2-year time points to suggest that any of these neoplastic findings were treatment-related and (2) the tumor morphologies and incidences were generally within the expected range of historical controls for Wistar Han rats. Findings from this study indicate that chronic exposure of male and female Wistar Han rats to either a tobacco blend used in snus, or a tobacco extract of that blend does not lead to increased toxicity or carcinogenicity, based on the specified outcomes measured.

Children under 15 kg with food allergy may be at risk of having epinephrine auto-injectors administered into bone.

BACKGROUND: The Epipen® Jr and Allerject® 0.15 mg are currently the most commonly prescribed epinephrine auto-injectors (EAIs) for the management of anaphylaxis in pediatric patients in North America and Canada. To ensure rapid absorption, it should be administered intramuscularly into the anterolateral aspect of the thigh. We examined whether the 12.7-mm needle length of the Epipen® Jr and Allerject® 0.15 mg is adequate for delivering epinephrine intramuscularly in pediatric patients who weighed <15 kg. METHODS: Consecutive pediatric patients with food allergy weighing <15 kg who required an EAI were included. Ultrasounds of the mid-anterolateral thigh were performed under minimal (min) and maximal (max) pressure. Skin-to-muscle depth (STMD) and skin-to-bone depth (STBD) measurements were completed. Baseline characteristics were compared between patients with a STBDmax <12.7 mm vs. ≥12.7 mm. Linear regression including variables such as age, sex, body mass index (BMI) and race was performed. The proportion of patients with a STBDmax <12.7 mm was compared in those weighing <10 kg vs. 10-14.9 kg. RESULTS: One hundred patients were included; 29 (29%) had STBDmax <12.7 mm. Height (p = 0.02) and weight (p = 0.0002) differed significantly between the two groups. Approximately 19% of those weighing 10-14.9 kg and 60% of those <10 kg had a STBDmax <12.7 mm. In the multivariable regression analysis, BMI was found to be a significant predictor of STBDmax. CONCLUSIONS: A large proportion of children <15 kg prescribed an EAI is at risk of having the auto-injector administered into bone. Since alternative EAIs with shorter needle lengths are not currently available, EAIs should be prescribed with appropriate counselling in this population.

Auto-injector needle length may be inadequate to deliver epinephrine intramuscularly in women with confirmed food allergy.

BACKGROUND: Epinephrine auto-injectors are the standard first aid treatment for anaphylaxis. Intramuscular delivery into the anterolateral aspect of the thigh is recommended for optimal onset of action of epinephrine. The most frequently prescribed auto-injector in North America and Canada is the EpiPen(®), which has a needle length of 15.2 mm. Currently, it is unknown whether this needle length is adequate for intramuscular delivery of epinephrine in adult patients at risk of anaphylaxis. METHODS: One hundred consecutive adult patients with confirmed food allergy requiring an epinephrine auto-injector were recruited. Skin to muscle depth (STMD) at the right mid-anterolateral thigh was measured using ultrasound under minimal (min) and maximum (max) pressure. The EpiPen(®) needle length was considered adequate if STMDmax was ≤15.2 mm. Baseline characteristics including age, gender, ethnicity, and body mass index (BMI) were compared in patients with STMDmax ≤15.2 mm vs. >15.2 mm. RESULTS: The EpiPen(®) needle length of 15.2 mm was inadequate for intramuscular delivery in 19 of the 100 enrolled patients (19%), all of whom were female; 28% of women had a STMDmax >15.2 mm. The mean STMDmax in the ≤15.2-mm and >15.2-mm groups were 9 ± 4 mm and 20 ± 4 mm, respectively (p = 0.0001). Linear regression analysis found BMI to be significantly associated with STMDmax after adjusting for age (p < 0.001). CONCLUSIONS: The needle length of the epinephrine auto-injectors may not be adequate for intramuscular delivery of epinephrine in a large proportion of women with food allergy; this may impact morbidity and mortality from anaphylaxis in this patient population.

Patients on subcutaneous allergen immunotherapy are at risk of intramuscular injections.

BACKGROUND: Allergen-specific subcutaneous immunotherapy is an effective treatment for certain allergic disorders. Ideally, it should be administered into the subcutaneous space in the mid-posterolateral upper arm. Injections are commonly given using a standard allergy syringe with a needle length of 13 mm. Therefore, there is a risk of intramuscular administration if patients have a skin-to-muscle depth <13 mm, which may increase the risk of anaphylaxis. The objective of this study was to determine whether the needle length of a standard allergy syringe is appropriate for patients receiving subcutaneous immunotherapy. METHODS: Ultrasounds of the left posterolateral arm were performed to measure skin-to-muscle depth in 200 adults receiving subcutaneous immunotherapy. The proportion of patients with a skin-to-muscle depth >13 mm vs. ≤13 mm was assessed and baseline characteristics of the two groups were compared. The proportion of patients with skin-to-muscle depths > 4 mm, 6 mm, 8 mm and 10 mm were also calculated. Multivariable logistic regression was performed to identify predictors of skin-to-muscle depth. RESULTS: Of the 200 patients included in the study, 80% had a skin-to-muscle depth ≤13 mm; the majority (91%) had a skin-to-muscle depth >4 mm. Body mass index was found to be a significant predictor of skin-to-muscle-depth. CONCLUSIONS: Most patients receiving subcutaneous immunotherapy have a skin-to-muscle depth less than the needle length of a standard allergy syringe (13 mm). These patients are at risk of receiving injections intramuscularly, which may increase the risk of anaphylaxis. Using a syringe with a needle length of 4 mm given at a 45° angle to the skin may decrease this risk.

Toxicological evaluation of smokeless tobacco: 90-day rodent feeding studies.

This manuscript presents data from 90-day toxicology studies designed to characterize the subchronic effects of a smokeless tobacco blend and an aqueous extract of that blend when administered to rodents in NTP-2000 feed. Positive control (nicotine tartrate) and treatment groups were matched for a range of nicotine levels. The doses evaluated were 0.3, 3, and 6 mg nicotine/kg body weight/day in Wistar Hannover rats and 6, 60, and 120 mg nicotine/kg/day in CD-1 mice. Variables evaluated included plasma nicotine and cotinine, body weights, feed consumption, clinical observations, clinical and anatomic pathology (including organ weights), and histopathology. Plasma nicotine and cotinine levels were dose-responsive. Key effects such as body weight reductions and organ weight changes occurred in rats and mice predominantly at the highest doses of test articles and positive control in the absence of treatment-related gross or histopathological changes. Organ weight changes were attributed mainly to the lower body weights of treated vs. control groups. The blend- and extract-induced effects generally paralleled each other and the nicotine-induced effects. Based on these studies, the doses evaluated spanned the no observable adverse effect level, the lowest observable adverse effect level and the maximum tolerated dose.

A Bayesian statistical analysis of mouse dermal tumor promotion assay data for evaluating cigarette smoke condensate.

The mouse dermal assay has long been used to assess the dermal tumorigenicity of cigarette smoke condensate (CSC). This mouse skin model has been developed for use in carcinogenicity testing utilizing the SENCAR mouse as the standard strain. Though the model has limitations, it remains as the most relevant method available to study the dermal tumor promoting potential of mainstream cigarette smoke. In the typical SENCAR mouse CSC bioassay, CSC is applied for 29 weeks following the application of a tumor initiator such as 7,12-dimethylbenz[a]anthracene (DMBA). Several endpoints are considered for analysis including: the percentage of animals with at least one mass, latency, and number of masses per animal. In this paper, a relatively straightforward analytic model and procedure is presented for analyzing the time course of the incidence of masses. The procedure considered here takes advantage of Bayesian statistical techniques, which provide powerful methods for model fitting and simulation. Two datasets are analyzed to illustrate how the model fits the data, how well the model may perform in predicting data from such trials, and how the model may be used as a decision tool when comparing the dermal tumorigenicity of cigarette smoke condensate from multiple cigarette types. The analysis presented here was developed as a statistical decision tool for differentiating between two or more prototype products based on the dermal tumorigenicity.

A summary of toxicological and chemical data relevant to the evaluation of cast sheet tobacco.

A tiered testing strategy based on a comparative chemical and biological testing program has been developed to evaluate the potential of tobacco processes, ingredients, or other technological developments to change the biological activity that results from burning tobacco. Cast sheet tobacco is a specific type of reconstituted tobacco sheet that can be used in the manufacture of cigarettes. The comparative chemical and biological testing program was used to compare the mainstream smoke and cigarette smoke condensate (CSC) from a Reference cigarette that did not contain cast sheet to that collected from Test cigarettes containing cast sheet at a final blend level of either 10% or 15%. Testing included mainstream cigarette smoke chemistry studies, in vitro studies (Ames assay, sister chromatid exchange assay, and neutral red cytotoxicity assay), and in vivo toxicology studies (13-week rat nose-only inhalation assay and 30-week mouse dermal tumor promotion assay). Certain statistically significant differences were observed in the chemical and biological studies when the Reference cigarette was compared to each of the Test cigarettes. However, when viewed collectively, the chemical and biological studies demonstrated that inclusion of cast sheet up to 15% in the final blend did not increase the inherent biological activity of mainstream cigarette smoke or CSC.

Upper airways sensory irritation responses of mice exposed to mainstream smoke from four cigarette types.

Relative sensory irritation responses for Swiss-Webster mice exposed nose-only to mainstream tobacco smoke were evaluated for several cigarette types using a smoking regimen consisting of a 35-ml puff, 2 s in duration, taken once per minute. The degree of sensory irritation for each cigarette type was evaluated as the smoke concentration inducing a 50% reduction in breathing frequency. The smoke concentration inducing 50% respiratory depression is called the RD(50) value. Study findings suggest that mainstream tobacco smoke from the Eclipse cigarette, which primarily heats rather than burns tobacco, yielded an RD(50) that was significantly higher (approximately twofold) than a tobacco-burning leading ultralight or the 2R4F or 1R5F reference cigarettes. This is indicative of reduced upper airways irritation by Eclipse that may be due to its distinct design. Study findings suggest that the irritating nature of mainstream tobacco smoke from different cigarette types can be evaluated effectively in terms of smoke concentration using the relative sensory irritation assessment. These findings constitute the first report about use of the RD(50) sensory irritation response during comparative evaluations of mainstream tobacco smoke.

Comparative 13-week inhalation study of cigarette smoke from cigarettes containing cast sheet tobacco.

A subchronic, nose-only inhalation study was conducted to compare the effects of mainstream smoke from a reference cigarette containing conventional reconstituted tobacco sheet at 30% of the finished blend to mainstream smoke from cigarettes containing 10% or 15% cast sheet (a specific type of reconstituted tobacco sheet) substituted for part of the conventional reconstituted tobacco. Male and female Sprague-Dawley rats were exposed for 1 h/day, 5 d/wk, for 13 wk to mainstream smoke at 0, 0.06, 0.20, or 0.80 mg wet total particulate matter per liter of air. Clinical signs, body and organ weights, clinical chemistry, hematology, carboxyhemoglobin (COHb), serum nicotine, plethysmography, gross pathology, and histopathology were determined. Exposure to cigarette smoke induced a number of changes in respiratory physiology, histopathology, and serum nicotine and COHb levels when compared to sham animals. When corresponding dose groups of reference and cast sheet mainstream smokes were compared, no biological differences were noted. At the end of the exposure period, subsets of rats from each group were maintained without smoke exposures for an additional 13 wk (recovery period). At the end of the recovery period, there were no statistically significant differences in histopathological findings observed between the reference and either cast sheet cigarette. Substitution of 10% or 15% cast sheet tobacco for conventional reconstituted tobacco sheet does not alter the inhalation toxicology of the mainstream smoke when compared to mainstream smoke from a reference cigarette containing conventional reconstituted tobacco sheet.

Cadmium exposure down-regulates 8-oxoguanine DNA glycosylase expression in rat lung and alveolar epithelial cells.

The current study tested the hypothesis that the pulmonary carcinogenic potential of cadmium (Cd) is related to its ability to inhibit the expression (mRNA and protein) and activity of 8-oxoguanine-DNA glycosylase (OGG1), a base excision repair (BER) enzyme that functions to preferentially excise pre-mutagenic 7,8-dihydro-8-oxoguanine (8-oxoG) from DNA. We demonstrate that a single Cd aerosol exposure of adult male Lewis rats causes time- and dose-dependent down-regulation in the pulmonary levels of rOGG1 mRNA and OGG1 protein, quantified by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assays and western analyses, respectively. Immunohistochemical studies confirmed that Cd inhalation reduces the relative amount of OGG1 in lungs of exposed animals without altering its over-all distribution within the lung, which appears to be more prominent within the alveolar epithelium. In agreement with our in vivo studies, we show that OGG1 expression is also attenuated in alveolar epithelial cell cultures exposed to CdCl(2) either acutely or by repeated passaging in Cd-containing medium. The effects caused by Cd were observed in cells that show no loss in viability, as assessed by colony forming ability, the MTT assay, and propidium iodide membrane permeability studies. Nuclear extracts prepared from Cd-treated cells also exhibit a reduction in the ability to nick a synthetic oligonucleotide containing 8-oxoG. We conclude from these studies that Cd causes suppression of OGG1 in the lung and that this mechanism may, in part, play a role in the Cd carcinogenic process.