INTRODUCTION
Inflammatory response is a biological process which involves response of the immune system.[1] This can be stimulated by pathogens, damaged cells, and toxic compounds.[1] These triggers may induce acute and/or chronic inflammatory responses in the body organs such as heart, pancreas, liver, kidney, lung, brain, intestinal tract, and reproductive system, which can lead to tissue damage or disease.[1]
Hypoglycemia can be referred to as low blood glucose which is a fall in blood sugar levels below the normal which can be caused by kidney failure, certain tumors (like insulinoma), liver disease, hypothyroidism, starvation, inborn error of metabolism, severe infections, reactive hypoglycemia, and drugs including alcohol. It is not a disease condition.[2,3,4,5]
Tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine involved in systemic inflammation and is one of the cytokines that constitutes the acute-phase reaction.[6,7] It is produced majorly by activated macrophages and by many other cell types such as CD4+ lymphocytes, NK cells, neutrophils, mast cells, eosinophils, and neurons.[6,7] TNF-α can induce insulin resistance by promoting serine phosphorylation of insulin receptor substrate-1 (IRS-1) that impairs insulin signaling.[6,7] Interleukin 10 (IL-10) or human cytokine synthesis inhibitory factor is an anti-inflammatory cytokine that can inhibit the synthesis or act against TNF-α.[6,7]
Diabetes mellitus (DM), commonly known as diabetes, is a chronic, metabolic disease characterized by elevated blood glucose leads which can lead to complications such as diabetic ketoacidosis, hyperosmolar hyperglycemic state or death, cardiovascular disease, stroke, chronic kidney disease, foot ulcers, and damage to the eyes.[8,9,10] Types of DM include Type 1 diabetes, (or juvenile diabetes or insulin-dependent diabetes,) which is a chronic condition in which the pancreas produces little or no insulin.[8,9,10] Individuals under this condition will depend on insulin treatment. The other type is the most commonly known as type 2 diabetes, usually in adults, which occurs when the body becomes resistant to insulin or does not make enough insulin.[8,9,10]
Insulin is a peptide hormone produced by beta cells of the islets of Langerhan of the pancreas to regulate metabolism of carbohydrates, fats, and protein by promoting the absorption of carbohydrates, especially glucose from the blood into liver, fat, and skeletal muscle cells.[11,12,13] Overdose or misapplication of insulin in DMis a common cause of death among diabetic patients.[11,12,13]
This work was designed to determine inflammatory responses in insulin-induced hypoglycemia among diabetes patients to provide information for useful guide in the management of diabetes mellitus.
METHODS
Ethical Consideration
The proposal of this work was presented, reviewed and approved Federal Medical Centre in Owo-Nigeria. Owo – Nigeria Consent of each subjects was also obtained.
Study area
The patients and controls were recruited through the Federal Medical Centre in Owo-Nigeria. Owo in Ondo state is an ancient Yoruba city located in the Southwestern part of Nigeria. It is equidistant to Lagos and Abuja (the Capital city of Nigeria). The city is a major link to South-South and North Central geopolitical zone in Nigeria. It is the headquarters of Owo local government in Owo-Ose Federal constituency. It hosts Achievers University and Rufus Giwa Polytechnic.
Study population
The study population include insulin-induced hypoglycemia diabetes patient (n = 30; female – 14; male – 16; 48–73 years), newly diagnosed diabetic patient (n = 30; female – 17; male – 13; 48–73 years) and nondiabetic patient (control (n = 50; female – 25; male – 25; 48–73 years) who were negative to Giemsa thick film technique for plasmodium, anti-hepatitis C virus (HCV), hepatitis B surface antigen (HBsAg), acid-fast bacilli, and Ag-Ab HIV tests.
Biological samples
Fasting venous blood was obtained from each of the participants.
Method of data analysis
The results of this work were subjected to statistical analysis using IBM SPSS 20.0 to determine the Student's ”t” value and probability at 0.05 level of significance (IBM, New York, USA).
Ethical consideration
The proposal of this study was reviewed and approved by the Research and Ethical Committee of the Federal Medical Centre, Owo, Nigeria. Informed consent was also obtained from each of the participants before the commencement of the work.
Methods of analysis of biological parameters
HIVp24 antigen and antibodies to HIV-1 (Groups M and O) and HIV-2 in human serum
These were determined in the participants using Bio-Rad Genscreen™ ULTRA HIV Ag-Ab qualitative enzyme immunoassay kit designed for the detection of HIV p24 antigen and antibodies to HIV-1 and HIV-2 in human serum/plasma by enzyme-linked immunosorbent assay (ELISA).
Principle
This ELISA method is based on the principle of the sandwich technique for the detection of HIV antigen and of the various antibodies associated with HIV-1 and/or HIV-2 virus in human serum or plasma. The solid phase is coated with monoclonal antibodies against p24 HIV-1 antigen, purified gp160 recombinant protein, epitope of HIV-1 group O-specific, and a peptide reacting like the epitope of the HIV-2 envelope protein. The test serum, control, and standard samples were added separately to the coated solid surface, incubated, washed, and allowed to react with the conjugate to give an observable product in the form of colored solution.
Antihepatitis C virus enzyme-linked immunosorbent assay
Anti-HCV was determined in the participants using Bio-Rad Monolisa™ Anti-HCV PLUS Version 3 screening kit designed for the detection of anti-HCV antibodies in human plasma or serum by ELISA technique (Bio-Rad Laboratories Company Hercules, California, United States).
Detection of hepatitis B surface antigen by enzyme-linked immunosorbent assay
HBsAg test was determined in the test and control volunteers by a one-step MONOLISA AgHBs PLUS enzyme immunoassay technique of the sandwich type for the detection of HBsAg in serum or plasma using the reagent kit of BIO-RAD Raymond Poincare, Marnes La Coquette (Bio-Rad Laboratories Company Hercules, California, United States).
Measurement of blood glucose
Blood glucose was determined in the participants by glucose oxidase method using the reagent kit of RANDOX (Crumlin, United Kingdom).
Tumor necrosis factor alpha enzyme-linked immunosorbent assay
Plasma TNF-α was analyzed using Abcam's TNF-α Human in vitro ELISA kit (Abcam, Cambridge, United Kingdom).
Interleukin-10 enzyme-linked immunosorbent assay
Plasma IL-10 was analyzed using Abcam's kit. Abcam's IL-10 Humanin vitro ELISA kit is designed for the quantitative measurement of IL-10 in supernatants, buffered solutions, and serum and plasma samples.
Detection of acid-fast bacilli in sputum and identification of plasmodium in blood
Acid-fast bacilli test and identification of plasmodium using sputum and blood samples, respectively, were carried out by the method, as described by Cheesbrough.[14]
RESULTS
The results obtained showed a significantly higher TNF-α and lower IL-10 in insulin-induced hypoglycemia diabetic patient than the results obtained in the newly diagnosed diabetic patient and nondiabetic participants [P < 0.05; Tables 1 and 2 and Figures 1 and 2].
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Table 1: Mean and standard deviation of plasma tumor necrosis factor-α, interleukin-10, fasting blood glucose, and alanine transaminase obtained in the participants (n=30)
Table 2: Comparative analysis of plasma tumor necrosis factor-α, interleukin-10, fasting blood glucose, and alanine transaminase obtained in the participants
Figure 1: Comparative description of plasma tumor necrosis factor-α, interleukin-10 and alanine aminotransferase in the participants
Figure 2: Comparative description of the values of fasting blood glucose in the participants
The results obtained showed a significantly higher TNF-α and lower IL-10 in a newly diagnosed diabetic patient than in nondiabetic participants [P < 0.05; Tables 1 and 2 and Figures 1 and 2].
The results obtained showed a significantly higher fasting blood glucose in a newly diagnosed diabetic patient than in nondiabetic participants [P < 0.05; Tables 1 and 2 and Figures 1 and 2].
The results obtained showed a significantly lower fasting blood glucose in insulin-induced hypoglycemia diabetic patient than the results obtained in the newly diagnosed diabetic patient and nondiabetic participants [P < 0.05; Tables 1 and 2 and Figures 1 and 2].
There was no significant difference in the results of plasma alanine transaminase obtained in the participants [P > 0.05; Tables 1 and 2 and Figures 1 and 2].
DISCUSSION
The results obtained showed a significantly higher TNF-α and lower IL-10 in insulin-induced hypoglycemia diabetic patient than the results obtained in the newly diagnosed diabetic patient and nondiabetic participants.
Increased TNF-α in insulin-induced hypoglycemia diabetes patients is attributable to its bioactivities of inducing insulin resistance in therapeutic misapplication of insulin in the treatment of diabetes as it will promote serine phosphorylation of IRS-1, to impair insulin signaling.[15] A common cause of hypoglycemia and death in diabetes patients is insulin overdose or misapplication.[16]
Insulin decreases blood glucose concentration by inducing cellular uptake of glucose because insulin causes the insertion of the GLUT4 transporter in the cell membranes of muscle and fat tissues which enables glucose to enter the cell.[12,17,18,19] It inhibits lipolysis, gluconeogenesis, and glycogenolysis.[12,17,18,19] Increase in these activities can result into hypoglycemia. Overdose or inappropriate administration of insulin in the treatment of diabetes can cause hypoglycemia resulting into inflammatory response with the release of TNF-α to induce insulin resistance.[12,17,18,19]
Decrease in IL-10 obtained in this study may be as a result of the increase in plasma TNF-α to regulate the activities of TNF-α because IL-10 is an anti-inflammatory cytokine, whereas TNF-α is a pro-inflammatory cytokine. IL-10 can inhibit the synthesis of pro-inflammatory cytokines such as IFN-γ, IL-2, IL-3, TNF-α, and GM-CSF.[20]
The results obtained showed a significantly higher TNF-α and lower IL-10 in newly diagnosed diabetic patient than in nondiabetic participants.
Increase in plasma TNF-α in diabetes patients can be due to the fact that several inflammatory cytokines are induced by oxidative stress.[21,22,23,24,25] Diabetes has been reported to be associated oxidative stress.[26,27] Furthermore, there is an immunological relationship between oxidative stress and cytokines because cytokines can trigger the release of other cytokines which can lead to increased oxidative stress.[21,22,23,24,25]
In addition, inflammation can also be induced by stress and chemicals such as toxin and drugs which can also be associated with the findings of this work.[28,29]
The results obtained showed a significantly higher fasting blood glucose in newly diagnosed diabetic patient than in nondiabetic participants. This indicates hyperglycemia which is a major characteristic of diabetes.[12] The results obtained showed a significantly lower fasting blood glucose in insulin-induced hypoglycemia diabetic patient than the results obtained in the newly diagnosed diabetic patient and nondiabetic participants. This indicates hypoglycemia induced by insulin.[12]
CONCLUSION
This work revealed significant inflammatory responses in insulin-induced hypoglycemia and diabetes as evidenced by increased plasma TNF-α and decreased plasma IL-10, which was more intense in insulin-induced hypoglycemia diabetes patients.
Financial support and sponsorship
Nil.
Conflicts of interest
There are no conflicts of interest.
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