Javascript is currently disabled in your browser. Several features of this site will not function whilst javascript is disabled.
open access to scientific and medical research
Papers Published
Open access peer-reviewed scientific and medical journals.
Learn more
Dove Medical Press is a member of the OAI.
Learn more
Bulk reprints for the pharmaceutical industry.
Learn more
We offer real benefits to our authors, including fast-track processing of papers.
Learn more
Register your specific details and specific drugs of interest and we will match the information you provide to articles from our extensive database and email PDF copies to you promptly.
Learn more
Back to Journals » Clinical Epidemiology » Volume 14
Authors Huang CC , Chen TH, Ho CH , Chen YC , Chen RJ, Wang YJ, Hsu CC, Lin HJ, Wang JJ, Chang CP, Guo HR
Received 6 July 2022
Accepted for publication 23 October 2022
Published 1 November 2022 Volume 2022:14 Pages 1265—1279
DOI https://doi.org/10.2147/CLEP.S380990
Checked for plagiarism Yes
Review by Single anonymous peer review
Peer reviewer comments 2
Editor who approved publication: Dr Lars Pedersen
Chien-Cheng Huang,1– 3 Tzu-Hao Chen,3,4 Chung-Han Ho,4,5 Yi-Chen Chen,4 Rong-Jane Chen,3,6 Ying-Jan Wang,3 Chien-Chin Hsu,1 Hung-Jung Lin,1,7 Jhi-Joung Wang,8,9 Ching-Ping Chang,4 How-Ran Guo3,10,11
1Department of Emergency Medicine, Chi Mei Medical Center, Tainan, Taiwan; 2Department of Emergency Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; 3Department of Environmental and Occupational Health, College of Medicine, National Cheng Kung University, Tainan, Taiwan; 4Department of Medical Research, Chi Mei Medical Center, Tainan, Taiwan; 5Department of Information Management, Southern Taiwan University of Science and Technology, Tainan, Taiwan; 6Department of Food Safety/Hygiene and Risk Management, College of Medicine, National Cheng Kung University, Tainan, Taiwan; 7Department of Emergency Medicine, Taipei Medical University, Taipei, Taiwan; 8Department of Anesthesiology, Chi Mei Medical Center, Tainan, Taiwan; 9Department of Anesthesiology, National Defense Medical Center, Taipei, Taiwan; 10Department of Occupational and Environmental Medicine, National Cheng Kung University Hospital, Tainan, Taiwan; 11Occupational Safety, Health and Medicine Research Center, National Cheng Kung University Hospital, Tainan, Taiwan
Correspondence: Chien-Cheng Huang, Department of Emergency Medicine, Chi Mei Medical Center, 901 Zhonghua Road, Yongkang District, Tainan, 710, Taiwan, Tel +886-6-281-2811, Fax +886-6-281-6161, Email [email protected] How-Ran Guo, Department of Environmental and Occupational Health, College of Medicine, National Cheng Kung University, 1 Daxue Road, Tainan, 701, Taiwan, Tel +886-6-235-3535, Fax +886-6-275-2484, Email [email protected]
Purpose: Carbon monoxide (CO) poisoning may damage the pancreas, but the effects of CO poisoning on the development of diabetes and on existing diabetes remain unclear. We conducted a study incorporating data from epidemiologic analyses and animal experiments to clarify these issues.
Methods: Using the National Health Insurance Database of Taiwan, we identified CO poisoning patients diagnosed between 2002 and 2016 (CO poisoning cohort) together with references without CO poisoning who were matched by age, sex, and index date at a 1:3 ratio. We followed participants until 2017 and compared the risks of diabetes and hyperglycemic crisis between two cohorts using Cox proportional hazards regressions. In addition, a rat model was used to assess glucose and insulin levels in blood as well as pathological changes in the pancreas and hypothalamus following CO poisoning.
Results: Among participants without diabetes history, 29,141 in the CO poisoning cohort had a higher risk for developing diabetes than the 87,423 in the comparison cohort after adjusting for potential confounders (adjusted hazard ratio [AHR]=1.23; 95% confidence interval [CI]: 1.18– 1.28). Among participants with diabetes history, 2302 in the CO poisoning cohort had a higher risk for developing hyperglycemic crisis than the 6906 in participants without CO poisoning (AHR = 2.12; 95% CI: 1.52– 2.96). In the rat model, CO poisoning led to increased glucose and decreased insulin in blood and damages to pancreas and hypothalamus.
Conclusion: Our epidemiological study revealed that CO poisoning increased the risks of diabetes and hyperglycemic crisis, which might be attributable to damages in the pancreas and hypothalamus as shown in the animal experiments.
Keywords: animal, carbon monoxide poisoning, diabetes, epidemiology, hypothalamus, pancreas
Carbon monoxide (CO) is a colorless, tasteless, and odorless gas produced by incomplete combustion of carbon compounds, including fire, engine exhaust, faulty furnaces, and charcoal-burning.1 CO poisoning is an important public health issue worldwide. In the United States, CO poisoning contributes to about 15,000 intentional poisoning and 50,000 emergency department visits annually, leading to 1319 deaths in 2014 alone.1 In Taiwan, a total of 25,912 CO poisoning cases between 1999 and 2012 were diagnosed, with a 1-month mortality rate of 1.6% and a 3-month mortality rate of 5.0%.2 In Asia, CO poisoning by charcoal-burning is a common method for suicide.3 In a study conducted in a medical center in Southern Taiwan, suicidal attempts accounted for 91.4% of CO poisoning cases.3 Another study in Northern Taiwan showed that 49.4% of CO poisoning cases were suicide attempts.4 In 2005, Taiwan government set up Taiwan Suicide Prevention Center to prevent suicide, including charcoal burning, which contributes to decreased percentage of suicide attempt in CO poisoning since 2007.2
The most well-known toxicity of CO is hypoxic injury, which is due to its high affinity to hemoglobin (Hb), about 250-fold of that of oxygen.1,5 Other mechanisms may involve inflammation, including mitochondrial dysfunction, increased oxidative stress and reactive oxygen species (ROS), etc.1,5 Neurological and myocardial injuries are the most common sequelae following CO poisoning due to the high oxygen demands of the brain and heart.1,5 Nationwide studies in Taiwan reported 6.2% incidence of neurological sequelae during a 12-month follow-up and 0.02% incidence of myocardial injury within 1 month of follow-up.2,6,7 However, CO toxicity may be systemic, not limited to the brain and heart. A nationwide study revealed that CO poisoning was associated with an increased risk of hypothyroidism and argued that injury to the hypothalamic-pituitary-thyroid axis was the cause.8 Recently, Weaver et al used a clinical dataset to investigate long-term health consequences of CO poisoning.9,10 They found that the incidence rates of hypothyroidism, bowel disease, and autoimmune disease, as well as later mortality and hospitalization were higher in patients with CO poisoning compared to the controls.9,10 Glucose homeostasis was regulated by the brain-endocrine pancreas axis, involving glucose-excited and glucose-inhibited neurons mainly located in the brainstem and hypothalamus, as well as β-cells in the pancreas.11 CO poisoning may lead to injuries to the brain and pancreas, and thus may increase the risk of diabetes. A nationwide study in Taiwan observed an increased risk of diabetes following CO poisoning.12 However, no epidemiologic or laboratory studies were conducted to verify the finding, despite that injury to the brain-endocrine pancreas axis is a plausible mechanism. Furthermore, the impact of CO poisoning on existing diabetes has not been evaluated. Therefore, we conducted a study incorporating larger epidemiologic data with an animal model to clarify these issues.
The Taiwan National Health Insurance Research Database (NHIRD) was used for the epidemiologic study.13 The NHIRD is generated from Taiwan National Health Insurance, which enrolled nearly all Taiwan’s citizens and provided a population-level evidence for clinical decisions and health policy making.13 It has also become an important basis for developing artificial intelligence in medical care in the future due to the “big data” feature.13
The study cohort (CO poisoning cohort) consisted of patients with CO poisoning who were identified from the NHIRD using International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) codes of 986, E868, E952, or E982 or International Statistical Classification of Diseases and Related Health Problems 10th Revision (ICD-10) code of T58 at the time of either hospitalization or emergency department care between 2002 and 2016.2,6–8,12,14 The index date was defined as the date when the patient was diagnosed with CO poisoning. In the study of the risk for developing diabetes, members of the CO poisoning cohort who had diabetes history were excluded, and a comparison cohort was then constructed by sampling candidates without diabetes history from the NHIRD who were matched with the non-diabetic CO poisoning cohort members by age, sex, and index date at a 3:1 ratio.6 Diabetes history was defined as presence of ICD-9-CM code of 250 or ICD-10 of E08-E13 in at least one hospitalization or three ambulatory care visits before the index date. Separate analyses were conducted on patients with diabetes history to evaluate the effects of CO poisoning on the diabetic population. Hyperglycemic crisis (ie, diabetic ketoacidosis or hyperosmolar hyperglycemic syndrome) was used as the indicator of the effects, and thus members of the CO poisoning cohort who had history of hyperglycemic crisis (ICD-9-CM of 250.1 or 250.2 or ICD-10 of E11.0 or E11.1) in any hospitalization were excluded. Likewise, a diabetic comparison cohort of members with diabetes history who did not have history of hyperglycemic crisis and were matched with the diabetic CO poisoning cohort members by age, sex, and index date at 1:3 ratio was constructed.
The diagnosis of CO poisoning is generally based on the following criteria: (1) confirmed exposure to CO, including elevated carboxyhemoglobin (COHb) levels or ambient CO concentrations, or a source of CO exposure (COHb level may not be elevated if there is a delay in obtaining the blood sample) and (2) presence of CO poisoning-related symptoms, including headache, dizziness, nausea, vomiting, malaise, fatigue, loss of consciousness, confusion, forgetfulness, visual disturbances, cardiac ischemia, and metabolic acidosis (a blood lactate level >2.5 mmol/L or a calculated base excess level <−2.0 mmol/L).15
Age was categorized as 18−34, 35−49, 50−64, and ≥65 years old.6 Underlying comorbidities were identified as the following: (1) hypertension: ICD-9-CM code of 401–405 or ICD-10 code of I10–I16; (2) obesity: ICD-9-CM code of 278 or ICD-10 code of E66; (3) smoking: ICD-9-CM code of 305.1 or ICD-10 code of Z72.0; (4) pre-diabetes: ICD-9-CM code of 790.2 or ICD-10 code of R73; (5) gestational diabetes: ICD-9-CM code of 648.0 or ICD-10 code of O24; (6) polycystic ovary syndrome: ICD-9-CM code of 256.4 or ICD-10 code of E28.2; (7) hyperlipidemia: ICD-9-CM code of 272 or ICD-10 code of E78.5; (8) hyperuricemia: ICD-9-CM code of 790.6 or 274 or ICD-10 code of E79.0; (9) chronic obstructive pulmonary disease: ICD-9-CM code of 490–496 or ICD-10 code of J44; (10) coronary artery disease: ICD-9-CM code of 410–414 or ICD-10 code of I20–I25; and (11) congestive heart failure: ICD-9-CM code of 428 or ICD-10 code of I50. They are the risk factors for diabetes and thus were potential confounders in the present study.16 Underlying comorbidities were defined as presence of these diagnostic codes in at least one hospitalization or three ambulatory care claims before the index date. This approach may minimize misclassification introduced by defining the outcome based on diagnostic codes on the claims.13
The outcomes of interest in the present study are diabetes and hyperglycemic crises. Cases of diabetes were defined as presence of ICD-9-CM code 250 or ICD-10 codes E08–E13 in at least one hospitalization or three ambulatory care visits after the index date. Cases of hyperglycemic crises were defined as presence of diabetic ketoacidosis or hyperosmolar hyperglycemic syndrome (ICD-9-CM code 250.1 or 250.2 or ICD-10 code E11.0 or E11.1) in at least one hospitalization after the index date.
Adult male Wistar rats (8–10 weeks old, weighing 380–420 g) purchased from the BioLASCO Taiwan Co., Ltd. (Taipei, Taiwan) were used for the study. We used only male rats because female rats have menstrual cycles, which may influence hormone modulation and some physiological parameters.17 The rats were housed in groups of four per cage in a temperature- and humidity-controlled animal facility with a 12-h light-dark cycle (08:00–20:00). Food and water were available ad libitum. We used 10–12 rats in each study group. All experiments involving experimental animals were conducted in accordance with the National Institutes of Health (NIH) guidelines for care and use of laboratory animals.
According to the protocol established in previous studies, we exposed the rats to 1000 ppm of CO for 20 min followed by 3000 ppm of CO for another 80 min in a 98-liter (38.1 cm × 50.8 cm × 50.8 cm) Plexiglas chamber.18 In the CO chamber, a constant flow of CO was set, and the concentration of CO was continuously monitored by a CO analyzer (TPI-708 carbon monoxide analyzer, Songdo-Dong, Yeonsu-Gu Incheon, Korea). We defined COHb ≥40% in the artery blood as the effective level for CO poisoning because it was the threshold of loss of consciousness (ie, acute severe CO poisoning) in human.19 The protocol (1000 ppm of CO in the air for 20 min followed by 3000 ppm of CO for another 80 min) was found to be the best after repetitive trials to achieve COHb of ≥40%.20 The rats were removed from the chamber to breathe fresh room air and regain consciousness after the above protocol of exposure.18 We removed all near-death rats and excluded them from data analyses. About 0.3 mL of artery blood from rat’s tail was drawn by PresetTM arterial blood collection syringe coating with heparin (364390, BD, Becton, USA), and the needle was replaced instead of 26 G 1/2 (301321, BD, Becton, USA). Once the blood was collected, sample was immediately preserved in ice bath and analyzed using blood gas analyzer (Nova Biomedical, Waltham, MA, USA) within 15 minutes to detect the level of COHb.
At the end of the experiment (day 28 after CO exposure, CO poisoning-Day 28), the rat was sacrificed, and the pancreas and brain tissues were harvested and subsequently fixed with formaldehyde solution (4%). After fixation, specimens were sequentially dehydrated with graded alcohol of 50% alcohol for 2 h, 70% alcohol for 2 h, 80% alcohol for 2 h, 95% alcohol for 1 h, 100% alcohol for overnight, and finally soaked in xylene for 90 minutes/time for 3 times. At last, specimens were embedded in paraffin, sliced into cuts of 3 μm by disposable microtome blades (08-635-0, ERMA, Kai industries CO., Ltd., Japan) on the micro slide glass (Pro-01, Matsunami, Japan), and stained using H&E staining. We examined pathological changes of the pancreas and hypothalamus using light microscopy and scored the damages according to a system used in previous studies.21–26
Specimens of the pancreas were processed using the same procedure as in H&E staining, including fixation and dehydration, for IF staining. We applied anti-insulin primary antibodies (ab6995, Abcam, USA) at a ratio of 1:100 and goat anti-mouse IgG secondary antibodies at a ratio of 1:400 (A11004, Invitrogen, USA) and then stained the specimens with 4’6-diamidino-2-phenylindole at a ratio of 1:10,000 (DAPI, Invitrogen, USA). After a final wash with phosphate buffered saline, sections were mounted in glycerol and examined using a confocal microscope with individual fluorescent wavelength (DAPI excitation/emission: 350–400/435–485 nm and TRITC excitation/emission:528–553/590–650 nm). We examined six 40× fields per section and recorded the average number of insulin-positive cells in each field.
Blood samples were drawn from the tail vein on the 1st, 7th, 14th, 21st, and 28th day after CO exposure. Serum samples were collected using centrifuge at the speed of 1500 × g for 15 min under 4-degree Celsius environment. We measured the level of glucose using a biochemical automatic analyzer (ABBOTT ARCHITECT c16000, Abbott Laboratories, Lake Forest, IL, USA) and the levels of insulin and glucagon using enzyme-linked immunosorbent assay (ELISA) kits (EZRMI-13K, Merck KGaA, Darmstadt, Germany and EZGLU-30K, Merck KGaA, Darmstadt, Germany, respectively) according to the manufacturer’s instructions.
The epidemiologic study was conducted according to the Declaration of Helsinki and after the approval of the Institutional Review Board (IRB) of the Chi Mei Medical Center. Patients’ informed consents were waived because this was a retrospective analysis of an anonymous database. The waiver did not affect the welfare of patients. Animal experiments were approved by the Institutional Animal Care and Use Committee (IACUC) of the Chi Mei Medical Center (IACUC No.108041101) and performed in accordance with the National Institutes of Health Guide for Care and Use of Laboratory Animals. The best effort was made to minimize animal suffering and number of animals sacrificed.
In the epidemiologic study, we used independent t-tests for continuous variables and chi-square tests for categorical variables to evaluate differences in demographic characteristics and underlying comorbidities between two cohorts. The Kaplan–Meier method with a Log rank test was performed to compare the risk of outcomes between two cohorts. We used multivariable Cox proportional hazards regressions to identify independent predictors of the outcomes and evaluate their effects. Stratified analyses according to age, sex, underlying comorbidities, and follow-up period were also conducted. All patients were right-censored to a new-onset outcome, date of death, or end of follow-up date, which is December 31, 2017. We used SAS 9.4 (SAS Institute Inc., Cary, NC, USA) to perform statistical analyses and STATA 14.0 (Stata Corp., College Station, TX, USA) to display Kaplan–Meier curves.
In the animal study, we used Prism (Version 7.01, S GraphPad Software, Inc., CA, USA) to perform statistical analyses. Data were presented as the mean ± standard error of the mean. Differences in pathophysiological scores between two groups of rats were evaluated using independent t-tests. Hormone values and glucose levels were analyzed using one-way analysis of variance for repeated measures. A two-tailed p value of <0.05 was considered statistically significant.
In non-diabetic CO poisoning and comparison cohorts, age subgroup of 18−34 years (44.8%) was the largest, followed by 35−49 years (36.7%), 50−64 years (13.7%), and ≥65 years (4.8%) (Table 1). The proportion of women was 51.7% in both cohorts because of matching. Compared to patients without CO poisoning, patients with CO poisoning had a higher prevalence of hypertension (7.1% vs 6.3%, p < 0.001), smoking (0.4% vs 0.2%, p < 0.001), chronic obstructive pulmonary disease (3.1% vs 1.9%, p < 0.001), coronary artery disease (2.2% vs 1.4%, p < 0.001), and congestive heart failure (0.6% vs 0.3%, p < 0.001).
 |
Table 1 Comparison of Age, Sex, and Underlying Comorbidities Between Patients with and without Carbon Monoxide (CO) Poisoning |
Table 1 Comparison of Age, Sex, and Underlying Comorbidities Between Patients with and without Carbon Monoxide (CO) Poisoning
Cox proportional hazards regressions showed that CO poisoning was an independent predictor for diabetes after adjustment for hypertension, obesity, smoking, pre-diabetes, polycystic ovary syndrome, hyperlipidemia, hyperuricemia, chronic obstructive pulmonary disease, coronary artery disease, and congestive heart failure (adjusted hazard ratio [AHR] = 1.23; 95% confidence interval [CI]: 1.18−1.28) (Table 2). In addition to CO poisoning, older age, male sex, hypertension, obesity, pre-diabetes, polycystic ovary syndrome, hyperlipidemia, hyperuricemia, chronic obstructive pulmonary disease, and congestive heart failure were also independent predictors for diabetes.
 |
Table 2 Independent Predictors for Diabetes Identified by Cox Proportional Hazards Regressions |
Table 2 Independent Predictors for Diabetes Identified by Cox Proportional Hazards Regressions
Stratified analyses showed that CO poisoning was associated with an increased risk of diabetes in all subgroups of age and sex, as well as those with underlying comorbidities of hypertension, hyperlipidemia, and hyperuricemia (Table 3). CO poisoning was associated with an increased risk of diabetes, and the AHR was 1.58 (95% CI: 1.42−1.77) in the first year, which decreased constantly afterwards but still reached statistical significance even after >6 years of follow-up (AHR = 1.15; 95% CI: 1.08−1.23). The Kaplan-Meier’s method and Log rank test also showed an increased risk of diabetes in the CO poisoning cohort (p < 0.001) (Figure 1).
 |
Table 3 Comparison of the Risk for Developing Diabetes Between Patients with and without Carbon Monoxide (CO) Poisoning Using Cox Proportional Hazards Regressions |
 |
Figure 1 Comparison of the risk for diabetes between patients with and without CO poisoning using Kaplan-Meier’s method and Log rank test. Abbreviation: CO, carbon monoxide. |
Table 3 Comparison of the Risk for Developing Diabetes Between Patients with and without Carbon Monoxide (CO) Poisoning Using Cox Proportional Hazards Regressions
Figure 1 Comparison of the risk for diabetes between patients with and without CO poisoning using Kaplan-Meier’s method and Log rank test.
Abbreviation: CO, carbon monoxide.
The CO poisoning cohort members with diabetes history had an increased risk of hyperglycemic crisis (AHR = 2.12; 95% CI: 1.52−2.96) compared with diabetic participants without CO poisoning (Table 4). In addition, stratified analyses showed an increased risk in patients with CO poisoning in subgroups of all age except age ≥65 years, both sexes, and with underlying hypertension. Moreover, the Kaplan-Meier’s method and Log rank test showed an increased risk of hyperglycemic crisis in the CO poisoning cohort members with diabetes compared to patients with diabetes but without CO poisoning (Figure 2).
 |
Table 4 Comparison of the Risk for Hyperglycemic Crises Between Diabetic Patients with and without Carbon Monoxide (CO) Poisoning Using Cox Proportional Hazards Regressions |
 |
Figure 2 Comparison of the risk for hyperglycemic crises between diabetic patients with and without CO poisoning using Kaplan-Meier’s method and Log rank test. Abbreviation: CO, carbon monoxide. |
Table 4 Comparison of the Risk for Hyperglycemic Crises Between Diabetic Patients with and without Carbon Monoxide (CO) Poisoning Using Cox Proportional Hazards Regressions
Figure 2 Comparison of the risk for hyperglycemic crises between diabetic patients with and without CO poisoning using Kaplan-Meier’s method and Log rank test.
Abbreviation: CO, carbon monoxide.
Compared to the sham group, CO poisoning rats (CO poisoning-Day 28) had more acinar cell dedifferentiation and necrosis, edema, loss of lobular integrity, parts of inflammation, and perivascular immune cell infiltration (Figure 3). CO poisoning rats showed structural disruption and vacuolation in the islet of Langerhans region (Figure 3A) and had higher damage scores in the comprehensive assessment (Figure 3B). The islet of Langerhans consists of five types of cells and secretes the following principal hormones: insulin (β-cells), glucagon (α-cells), somatostatin (δ-cells), pancreatic polypeptide (PP cells), and ghrelin (ε-cells).27 Insulin and glucagon are released directly into the blood circulation to regulate glucose levels.27 Because disruption of the pancreatic islet was observed, we examined the levels of glucose, insulin, and glucagon in the blood. CO poisoning rats had increased glucose levels after CO poisoning-Day 7 and decreased insulin levels on CO poisoning-Days 1, 7, 14, 21, and 28 (Figure 3C and D). However, differences in the glucagon level between the CO poisoning group after CO poisoning and the sham group did not reach statistical significance (Figure 3E). IF staining showed that the sham group had sufficient insulin in the pancreatic islets (Figure 4A), but CO poisoning rats had insufficient insulin production (Figure 4B). The average number of insulin-positive cells per field on CO poisoning-Day 28 in the CO poisoning group was less than that in the sham group (Figure 4C).
 |
Figure 3 Pathological change in the pancreas with different magnification using hematoxylin-eosin stain and relevance of biochemical parameters manifestation. Upper row represents the sham group and the bottom row represents the CO poisoning group on CO poisoning-Day 28 (A) and statistical quantification of damage scores between two groups (B), n = 10 per group, ****p < 0.0001. Biochemical indicator like glucose (C) and relative hormone like insulin (D) and glucagon (E) were detected from the serum by ELISA assay at the indicated day, n = 10–12 per group, **p < 0.01, ***p < 0.001, ****p < 0.0001. Abbreviation: CO, carbon monoxide. |
 |
Figure 4 Insulin expression by immunofluorescence staining. Pancreatic islet was labeled by insulin antibody co-staining with nucleus staining (DAPI) shown in different magnification (200×–400×) in the sham (A) and CO poisoning group (B). Comparison of the number of insulin-positive cells per field was showed in the (C) ***p < 0.001. Abbreviation: CO, carbon monoxide. |
Figure 3 Pathological change in the pancreas with different magnification using hematoxylin-eosin stain and relevance of biochemical parameters manifestation. Upper row represents the sham group and the bottom row represents the CO poisoning group on CO poisoning-Day 28 (A) and statistical quantification of damage scores between two groups (B), n = 10 per group, ****p < 0.0001. Biochemical indicator like glucose (C) and relative hormone like insulin (D) and glucagon (E) were detected from the serum by ELISA assay at the indicated day, n = 10–12 per group, **p < 0.01, ***p < 0.001, ****p < 0.0001.
Abbreviation: CO, carbon monoxide.
Figure 4 Insulin expression by immunofluorescence staining. Pancreatic islet was labeled by insulin antibody co-staining with nucleus staining (DAPI) shown in different magnification (200×–400×) in the sham (A) and CO poisoning group (B). Comparison of the number of insulin-positive cells per field was showed in the (C) ***p < 0.001.
Abbreviation: CO, carbon monoxide.
The hypothalamus-pituitary gland axis is well known for its modulatory function of hormonal secretion. Corticotropin-releasing hormone, secreted by the hypothalamus, increases the release of insulin, indicating that functional receptor systems for hypothalamic-releasing hormone agonists exist within the endocrine pancreas.28 Therefore, we evaluated whether the hypothalamus, an upstream organ, can be injured after CO poisoning. Severe damages such as pyknosis, karyorrhexis, vacuolation, and immune cell infiltration in the hypothalamus of CO poisoning rats were observed in the pathophysiological assessment (Figure 5A). The damage score of the hypothalamus on CO poisoning-Day 28 in the CO poisoning group was higher than that in the sham group (Figure 5B).
 |
Figure 5 Histopathological change of the brain region by hematoxylin-eosin stain. Significant damages such as pyknosis, karyorrhexis, vacuolation, and immune cell infiltration were found in the hypothalamus of CO poisoning rats (A). The damage score of the hypothalamus was significantly higher in the CO poisoning-Day 28 group than that of the sham group (B). ***p < 0.001. Abbreviation: CO, carbon monoxide. |
Figure 5 Histopathological change of the brain region by hematoxylin-eosin stain. Significant damages such as pyknosis, karyorrhexis, vacuolation, and immune cell infiltration were found in the hypothalamus of CO poisoning rats (A). The damage score of the hypothalamus was significantly higher in the CO poisoning-Day 28 group than that of the sham group (B). ***p < 0.001.
Abbreviation: CO, carbon monoxide.
The epidemiologic study revealed that CO poisoning increased the risk of developing diabetes and increased the risk of hyperglycemic crisis in patients with diabetes. In the animal study, an increased level of glucose and a decreased level of insulin in the blood as well as significant damages in the pancreatic islets and hypothalamus were found, which could explain findings in the epidemiologic study. In addition to CO poisoning, we found that older age, male sex, hypertension, obesity, pre-diabetes, polycystic ovary syndrome, hyperlipidemia, hyperuricemia, chronic obstructive pulmonary disease, and congestive heart failure were independent predictors for diabetes, which is compatible with previous studies.16
The most likely cause of the increased risks is CO poisoning-related hypoxic injury, mainly in the beta cells of the pancreatic islets.29–31 Pancreatic beta cells have high oxygen demand during the process of insulin secretion, making pancreatic beta cells very sensitive to hypoxia.30 Previous studies showed that hypoxia during islet transplantation contributed to beta cell dysfunction and apoptosis via the mitochondrial cell death pathway.32,33 Hypoxia induces endoplasmic reticulum stress and upregulates the pro-apoptotic transcription factor C/EBP homologous protein, which initiates apoptotic cell death.30 Beta cell dysfunction and apoptosis may lead to decreased insulin secretion and subsequent hyperglycemia,31 which is compatible with the increased risk of developing diabetes in the non-diabetic population and the increased risk of hyperglycemic crisis in the diabetic population following CO poisoning observed in the present study.
The positive association between CO poisoning and diabetes observed in the present study may help explain the association between smoking and diabetes in previous studies.34,35 Smoking is a predictor for progression of glucose intolerance, including the progression from normoglycemia to impaired glucose tolerance status and the development of diabetes.34,35 One of the plausible mechanisms is that CO exposure during smoking leads to hypoxic injuries of the beta cells.29–31,35 Duration of smoking also has a positive association with the risk of diabetes, and this dose-response relationship suggests a causal link.35 Although the present study has provided evidence supporting that CO exposure plays a role in the association between smoking and diabetes, the underlying mechanism needs further studies to clarify, because many compounds are involved in smoking such as tar, arsenic, lead, and nicotine.35
Mitochondrial dysfunction, ROS generation, and inflammation may also be mechanisms through which CO poisoning increases the risk of diabetes. Patients with diabetes have elevated levels of inflammation markers and mediators, as well as acute-phase reactants (including tumor necrosis factor-alpha, fibrinogen, C-reactive protein, interleukin 6, plasminogen activator inhibitor-1, and sialic acid) and white blood cells.36 A human study by Chambers et al showed that CO severity did not predict 1 year outcome, supporting the idea that inflammation may be the etiology.37 A much less severe animal poisoning model would be very informative for this issue. Two major mechanisms related to mitochondrial dysfunction can affect the development and deterioration of diabetes: increased insulin resistance and decreased insulin secretion.38 Previous human and animal studies revealed impaired oxidative phosphorylation in muscle mitochondria in insulin-resistant states.39,40 A study using mitochondria isolated from human muscle biopsy specimens observed decreased nicotinamide adenine dinucleotide + hydrogen:O(2) oxidoreductase activity, citrate synthase activity, and smaller skeletal muscle mitochondria in patients with diabetes than in lean volunteers, and concluded that mitochondrial dysfunction in skeletal muscle is associated with diabetes.40 Mitochondrial dysfunction affects insulin secretion via impairing adenosine triphosphate production,38 which is essential for calcium entry and insulin release from storage granules, making it possible for mitochondrial dysfunction and excess ROS to cause oxidative damage and insulin secretion impairment.
The severe damages of the hypothalamus observed in the present study may also elucidate the mechanisms of increased diabetes following CO poisoning. Glucose homeostasis is controlled by glucagon, insulin, and autonomic nervous systems.11 Activation or inhabitation of glucose-sensing neurons in the hypothalamus may alter glucose homeostasis and contribute to the increased risk of diabetes and poor control of existing diabetes. Direct damage to the pancreas is another mechanism through which CO poisoning may lead to diabetes. A meta-analysis showed that 15% (95% CI: 9%−22%) of the patients with first episode of acute pancreatitis developed new-onset diabetes with 1 year and the relative risk still reached statistical significance after five years of follow up.44 In the present study, the AHR of diabetes associated CO poisoning was 1.58 (95% CI: 1.42−1.77) in the first year, which still reached statistical significance even after 6 years of follow-up. The findings are compatible with those in the meta-analysis.
Immunological responses following CO poisoning are another possible mechanism of the long-term health effects, including diabetes and other disorders.41,42 An animal study reported that CO poisoning contributes to adduct formation between myelin basic protein and malonylaldehyde, which causes an immunological cascade and plays a role in delayed neurological sequelae.41 Compared with the controls, a human study showed that an array of autoantibodies elevated in the patients with CO poisoning.42 A recent study by Weaver et al showed that autoantibodies and neural proteins were often elevated in the participants with CO poisoning compared with the controls.43 These studies shed some light on the mechanism and potential biomarker for predicting adverse outcome of CO poisoning.41–43
The major strength of the present study is the combination of epidemiologic and animal evidence. In addition, the large sample size of the epidemiologic study allowed adjustment of most potential confounders. The major limitation is the lack of information on some risk factors for diabetes which were not available in the NHIRD, including family history of diabetes and lifestyle such as diet and physical inactivity. However, the animal study, which was not affected by these factors that were not measured in the epidemiologic study, showed increased blood glucose level associated with damages of pancreatic islets and decreased insulin secretion after CO exposure. These findings supported the observations in the epidemiologic study. Second, our animal study may not be representative of CO poisoning in humans. Third, the patients were not coded for subgroups of diabetes, such as type 1, type 2, gestational, etc., and so we were not able to perform further analyses. Further studies to identify which subgroup is the most affected or which is the least affected by CO exposure are warranted.
The epidemiologic data showed that CO poisoning increased the risk of developing diabetes and the risk of hyperglycemic crisis (a sign of poor control of diabetes) in patients with existing diabetes. The most plausible cause is that CO poisoning contributes to hypoxic injuries to the beta cells of the pancreatic islets and subsequently decreases insulin secretion. Other causes, including mitochondrial dysfunction, ROS generation, inflammation, autoantibodies formation, and damage of the hypothalamus may also play a role. Therefore, close follow-up for the development of diabetes, or poor control of existing diabetes are recommended for patients with CO poisoning. The present study may also become an important reference for the research of low-level and long-term exposure of CO, such as those from air pollution, working environment, and tobacco smoke.
We thank Dr. Wei-Ting Chang for assistance to the animal study, and Enago for the English editing.
All authors made a significant contribution to the work reported, whether that is in the conception, study design, execution, acquisition of data, analysis and interpretation, or in all these areas; took part in drafting, revising or critically reviewing the article; gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agree to be accountable for all aspects of the work.
This study was supported by Grant MOHW108-TDU-B-212-124014 from the Ministry of Health and Welfare, Taiwan R.O.C. using surcharge of tobacco products, Grants MOST 111-2314-B-384-007, MOST 111-2314-B-006-031-MY3, MOST 109-2314-B-384-005-MY3 and MOST 108-2238-B-006-001-MY2 from the Ministry of Science and Technology, Taiwan, R.O.C., and Grant Physician-Scientist 11001 from the Chi Mei Medical Center.
No conflicts of interest to declare.
1. Rose JJ, Wang L, Xu Q, et al. Carbon monoxide poisoning: pathogenesis, management, and future directions of therapy. Am J Respir Crit Care Med. 2017;195(5):596–606. doi:10.1164/rccm.201606-1275CI
2. Huang CC, Ho CH, Chen YC, et al. Demographic and clinical characteristics of carbon monoxide poisoning: nationwide data between 1999 and 2012 in Taiwan. Scand J Trauma Resusc Emerg Med. 2017;25(1):70. doi:10.1186/s13049-017-0416-7
3. Huang CC, Lee JC, Lin KC, et al. Exposure duration and history of hypertension predicted neurological sequelae in patients with carbon monoxide poisoning. Epidemiology. 2019;30(Suppl 1):S76–S81. doi:10.1097/EDE.0000000000001000
4. Ku CH, Hung HM, Leong WC, et al. Outcome of patients with carbon monoxide poisoning at a far-east poison center. PLoS One. 2015;10(3):e0118995. doi:10.1371/journal.pone.0118995
5. Weaver LK. Clinical practice. Carbon monoxide poisoning. N Engl J Med. 2009;360(12):1217–1225. doi:10.1056/NEJMcp0808891
6. Huang -C-C, Ho C-H, Chen Y-C, et al. Risk of myocardial infarction after carbon monoxide poisoning: a nationwide population-based cohort study. Cardiovasc Toxicol. 2019;19(2):147–155. doi:10.1007/s12012-018-9484-9
7. Huang CC, Ho CH, Chen YC, et al. Effects of hyperbaric oxygen therapy on acute myocardial infarction following carbon monoxide poisoning. Cardiovasc Toxicol. 2020;20(3):291–300. doi:10.1007/s12012-019-09552-7
8. Huang CC, Ho CH, Chen YC, et al. Increased risk for hypothyroidism associated with carbon monoxide poisoning: a nationwide population-based cohort study. Sci Rep. 2019;9(1):16512. doi:10.1038/s41598-019-52844-9
9. Weaver LK, Churchill S, Calderone PE, Perez AC, Schneider M. Retrospective cohort study of thyroid, bowel, and autoimmune diseases after carbon monoxide poisoning. 2022 Annual Scientific Meeting of the Undersea and Hyperbaric Medical Society Meeting. Reno, NV, USA; 2022.
10. Deru KW, Churchill S, Calderone P. Retrospective case-control study of long-term health outcomes after carbon monoxide poisoning. Undersea Hyperb Med; 2021: 330.
11. Thorens B. Brain glucose sensing and neural regulation of insulin and glucagon secretion. Diabetes Obes Metab. 2011;13(Suppl 1):82–88. doi:10.1111/j.1463-1326.2011.01453.x
12. Huang CC, Ho CH, Chen YC, et al. Increased risk for diabetes mellitus in patients with carbon monoxide poisoning. Oncotarget. 2017;8(38):63680–63690. doi:10.18632/oncotarget.18887
13. Hsieh CY, Su CC, Shao SC, et al. Taiwan’s national health insurance research database: past and future. Clin Epidemiol. 2019;11:349–358. doi:10.2147/CLEP.S196293
14. Huang CC, Ho CH, Chen YC, et al. Hyperbaric oxygen therapy is associated with lower short- and long-term mortality in patients with carbon monoxide poisoning. Chest. 2017;152(5):943–953. doi:10.1016/j.chest.2017.03.049
15. Huang CC, Ho CH, Chen YC, et al. Impact of hyperbaric oxygen therapy on subsequent neurological sequelae following carbon monoxide poisoning. J Clin Med. 2018;7:32.
16. Robertson RP. Risk factors for type 2 diabetes mellitus. J Cardiovasc Nurs. 2020;2020:17–23.
17. Ajayi AF, Akhigbe RE. Staging of the estrous cycle and induction of estrus in experimental rodents: an update. Fertil Res Pract. 2020;6(1):5. doi:10.1186/s40738-020-00074-3
18. Fan DF, Hu HJ, Sun Q, et al. Neuroprotective effects of exogenous methane in a rat model of acute carbon monoxide poisoning. Brain Res. 2016;1633:62–72. doi:10.1016/j.brainres.2015.12.019
19. Hampson NB, Hauff NM. Carboxyhemoglobin levels in carbon monoxide poisoning: do they correlate with the clinical picture? Am J Emerg Med. 2008;26(6):665–669. doi:10.1016/j.ajem.2007.10.005
20. Huang CC, Chen TH, Ho CH, et al. Increased risk of congestive heart failure following carbon monoxide poisoning. Circ Heart Fail. 2021;14(4):e007267. doi:10.1161/CIRCHEARTFAILURE.120.007267
21. R Chen, Wang J, Liao C, Zhanga. L, Wang. X. 1H NMR studies on serum metabonomic changes over time in a kidney-Yang deficiency syndrome model. RSC Adv. 2017;54:34251–34261.
22. Folias AE, Penaranda C, Su AL, Bluestone JA, Hebrok M. Aberrant innate immune activation following tissue injury impairs pancreatic regeneration. PLoS One. 2014;9(7):e102125. doi:10.1371/journal.pone.0102125
23. Honorio JE Jr., Vasconcelos GS, Rodrigues FT, et al. Monocrotaline: histological damage and oxidant activity in brain areas of mice. Oxid Med Cell Longev. 2012;2012:697541. doi:10.1155/2012/697541
24. Liu X, Gu X, Li Z, et al. Neuregulin-1/erbB-activation improves cardiac function and survival in models of ischemic, dilated, and viral cardiomyopathy. J Am Coll Cardiol. 2006;48(7):1438–1447. doi:10.1016/j.jacc.2006.05.057
25. Schmidt J, Rattner DW, Lewandrowski K, et al. A better model of acute pancreatitis for evaluating therapy. Ann Surg. 1992;215(1):44–56. doi:10.1097/00000658-199201000-00007
26. Syro LV, Rotondo F, Cusimano MD, et al. Current status on histological classification in Cushing’s disease. Pituitary. 2015;18(2):217–224. doi:10.1007/s11102-014-0619-0
27. Zhou Q, Melton DA. Pancreas regeneration. Nature. 2018;557(7705):351–358. doi:10.1038/s41586-018-0088-0
28. Schmid J, Ludwig B, Schally AV, et al. Modulation of pancreatic islets-stress axis by hypothalamic releasing hormones and 11beta-hydroxysteroid dehydrogenase. Proc Natl Acad Sci U S A. 2011;108(33):13722–13727. doi:10.1073/pnas.1110965108
29. Gunton JE. Hypoxia-inducible factors and diabetes. J Clin Invest. 2020;130(10):5063–5073. doi:10.1172/JCI137556
30. Zheng X, Zheng X, Wang X, et al. Acute hypoxia induces apoptosis of pancreatic beta-cell by activation of the unfolded protein response and upregulation of CHOP. Cell Death Dis. 2012;3(6):e322. doi:10.1038/cddis.2012.66
31. Sherwani SI, Aldana C, Usmani S, et al. Intermittent hypoxia exacerbates pancreatic beta-cell dysfunction in a mouse model of diabetes mellitus. Sleep. 2013;36(12):1849–1858. doi:10.5665/sleep.3214
32. Olsson R, Olerud J, Pettersson U, Carlsson PO. Increased numbers of low-oxygenated pancreatic islets after intraportal islet transplantation. Diabetes. 2011;60(9):2350–2353. doi:10.2337/db09-0490
33. Lau J, Henriksnas J, Svensson J, Carlsson PO. Oxygenation of islets and its role in transplantation. Curr Opin Organ Transplant. 2009;14(6):688–693. doi:10.1097/MOT.0b013e32833239ff
34. Sliwinska-Mosson M, Milnerowicz H. The impact of smoking on the development of diabetes and its complications. Diab Vasc Dis Res. 2017;14(4):265–276. doi:10.1177/1479164117701876
35. Foy CG, Bell RA, Farmer DF, Goff DC Jr., Wagenknecht LE. Smoking and incidence of diabetes among U.S. adults: findings from the Insulin Resistance Atherosclerosis Study. Diabetes Care. 2005;28(10):2501–2507. doi:10.2337/diacare.28.10.2501
36. Tsalamandris S, Antonopoulos AS, Oikonomou E, et al. The role of inflammation in diabetes: current concepts and future perspectives. Eur Cardiol. 2019;14(1):50–59. doi:10.15420/ecr.2018.33.1
37. Chambers CA, Hopkins RO, Weaver LK, Key C. Cognitive and affective outcomes of more severe compared to less severe carbon monoxide poisoning. Brain Inj. 2008;22(5):387–395. doi:10.1080/02699050802008075
38. Sivitz WI, Yorek MA. Mitochondrial dysfunction in diabetes: from molecular mechanisms to functional significance and therapeutic opportunities. Antioxid Redox Signal. 2010;12(4):537–577. doi:10.1089/ars.2009.2531
39. Kelley DE, He J, Menshikova EV, Ritov VB. Dysfunction of mitochondria in human skeletal muscle in type 2 diabetes. Diabetes. 2002;51:2944–2950.
40. Boudina S, Sena S, Theobald H, et al. Mitochondrial energetics in the heart in obesity-related diabetes: direct evidence for increased uncoupled respiration and activation of uncoupling proteins. Diabetes. 2007;56(10):2457–2466. doi:10.2337/db07-0481
41. Thom SR, Bhopale VM, Fisher D, Zhang J, Gimotty P. Delayed neuropathology after carbon monoxide poisoning is immune-mediated. Proc Natl Acad Sci U S A. 2004;101(37):13660–13665. doi:10.1073/pnas.0405642101
42. Thom SR, Bhopale VM, Milovanova TM, et al. Plasma biomarkers in carbon monoxide poisoning. Clin Toxicol. 2010;48(1):47–56. doi:10.3109/15563650903468209
43. Weaver LK, Deru K, Churchill S, Carlquist J. Neural proteins and auto-antibodies after carbon monoxide poisoning. Undersea Hyperb Med. 2021;48:325–326.
44. Das SL, Singh PP, Phillips AR, et al. Newly diagnosed diabetes mellitus after acute pancreatitis: a systematic review and meta-analysis. Gut. 2014;63(5):818–831. doi:10.1136/gutjnl-2013-305062
© 2022 The Author(s). This work is published and licensed by Dove Medical Press Limited. The full terms of this license are available at https://www.dovepress.com/terms.php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License. By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms.
Contact Us • Privacy Policy • Associations & Partners • Testimonials • Terms & Conditions • Recommend this site • Top
Contact Us • Privacy Policy
© Copyright 2022 • Dove Press Ltd • software development by maffey.com • Web Design by Adhesion
The opinions expressed in all articles published here are those of the specific author(s), and do not necessarily reflect the views of Dove Medical Press Ltd or any of its employees.
Dove Medical Press is part of Taylor & Francis Group, the Academic Publishing Division of Informa PLC
Copyright 2017 Informa PLC. All rights reserved. This site is owned and operated by Informa PLC ( “Informa”) whose registered office is 5 Howick Place, London SW1P 1WG. Registered in England and Wales. Number 3099067. UK VAT Group: GB 365 4626 36
In order to provide our website visitors and registered users with a service tailored to their individual preferences we use cookies to analyse visitor traffic and personalise content. You can learn about our use of cookies by reading our Privacy Policy. We also retain data in relation to our visitors and registered users for internal purposes and for sharing information with our business partners. You can learn about what data of yours we retain, how it is processed, who it is shared with and your right to have your data deleted by reading our Privacy Policy.
If you agree to our use of cookies and the contents of our Privacy Policy please click ‘accept’.