| Introduction:Tuberculosis is a chronic disease caused by Mycobacterium tuberculosis.It has a history of more than seven thousand years.The studies of archaeologists based on the human bone fossils in Neolithic age as well as in the mummies of Egyptians 4500 years ago have found the spinal tuberculosis.Many organs can be invaded by Mycobacterium tuberculosis,especially tuberculosis,and it is one of the most important diseases that endanger human health.In 1993,because tuberculosis has become a major threat to global public health and has become a major disease and mortality increasing in many countries,the WHO declared that the global tuberculosis was at a state of emergency.According to the WHO global report in 2017,there were about 10,400,000 new tuberculosis cases occurred in 2015.In 2015,there were 918,000 new tuberculosis cases in China,including 296,000 females and 622,000 males.The fatality rate was 4%.In recent years,with the growth of the floating population,we have two major problems in the treatment of tuberculosis,that is,drug-resistant tuberculosis and tubercular patients with HIV infection.In 2015,China's data showed that about 57,000 patients with MDR-TB and RR-TB,10034 tubercular patients with positive HIV,while only 3750 of those were under treatment,accounting for only 37%.These two kinds of patients are not only expensive in treatment,but also poor in effect and easy to spread,which is the focus of tuberculosis control.China has a heavy burden of tuberculosis in the world,and ranking has been in the second place for a long time,only from second to third in 2015.On the one hand,it shows the importance and input of tuberculosis in China,and has certain effects.On the other hand,it shows the seriousness of tuberculosis in our country.In 2016,the cost of tuberculosis in China was about $37 million 200 thousand,or about 2 billion 500 million 800 thousand RMB.This is a huge economic burden for our country.Diabetes is a chronic disease that seriously threaten human health too.With the changes of people's lifestyle and the aging of the population structure,the incidence of diabetes is increasing rapidly in China and even in the world.The International Diabetes Federation(IDF)had announced the 7th IDF diabetes map in December 1,2015,which introduced the prevalence and economic burden of current global population of diabetes,ranging in age from 20 to 79,and the incidence of diabetes in the future trend of global forecast,and the incidence of diabetes in the future is predicted.In 2015,the number of people with diabetes in the world rose to 415 million,and the prevalence rose to 8.8%.75%of these patients are distributed in low and middle-income countries.The global prevalence of diabetes is on the rise,and the number of patients with diabetes is expected to reach 642 million in 2040,and the prevalence rate will rise to 10.4%(roughly 8.5%-13.5%).The number of patients with diabetes in China has ranked first in the world in 2015.The number of diabetic patients in China increased by 11 million 200 thousand compared with 2013,up to 109 million 600 thousand.According to the current trend of development,the population of diabetes in China is expected to reach 150 million 700 thousand by 2040.In 2015,the cost of diabetes treatment and prevention of complications in the world has amounted to $673-1197 billion,accounting for 11.6%of global medical expenditure.By 2040,the global healthcare costs associated with diabetes will reach$802-1452 billion.The diabetics are 3 times more likely to be infected with tuberculosis than the general population.Due to the disorders of glucose,protein and fat metabolism of diabetics,resulting in malnutrition,decrease the capacity of body's resistance and tissue repair.In the state of low immunity for long term,at the same time,the glucose in blood and tissue is increased,and with the accumulation of glycerol products,which is a good medium for bacteria,while microvascular lesions involving the lungs,also conducive to the growth and reproduction of mycobacterium.However,the blood glucose of tubercular patients with diabetes is often too high,thus it's difficult to control well.Therefore,the occurrence of diabetes and tuberculosis affect each other,and there is a reciprocal causation.And influence treatment effect of each other.In recent years,in the global tuberculosis control improvement trend,diabetes mellitus patients with tuberculosis are increasing year by year.In recent years,under the improved trend of global TUBERCULOSIS control,but the number of patients with diabetes and tuberculosis is increasing year by year.China ranks second in the countries with high rates of tuberculosis and diabetes.In 2012,data have showed that the diabetics account for 17%of adults with active pulmonary tuberculosis in China,and there were 156000 patients with tuberculosis and diabetes.Thus,to prevent and control tuberculosis,attention should be paid to the control of diabetes.At present,although we have a lot of tuberculosis diagnosis and monitoring methods,but there are deficiencies.Some of the methods still need a long time,such as Mycobacterium culture,still need more than a month,and plus the drug sensitive test,it will take more than 40 days.Before the results come out,we can only use the drugs according to the clinical experience.Chest image examination was often used to judge the effectiveness of treatment,but due to the body's radiation,it can not be checked frequently;and so on.It is difficult to meet the actual needs of the clinic.Therefore,we urgently need a method for diagnosing tuberculosis with high sensitivity and specificity.At present,the gene therapy has been researched in each field,and some of them have achieved success in some fields.However,there are still few studies on TB gene.MicroRNA(miRNA)is a kind of endogenous expression of small single stranded RNA genome,in non encoding region,plays an important role in cell growth,development,differentiation,death and other biological process,and the length is 18 to 25 nucleotides,which can be formed by the processing of hairpin shaped 60-110nt endogenous transcript precursor.Recent studies have shown that miRNA can be used as a biological marker for the early diagnosis of certain diseases,including stomach cancer,colorectal cancer,respiratory system diseases,cardiovascular diseases,and some prenatal diagnosis.Although it is short in length,it can affect the expression of many proteins and play a role in regulating gene expression during target gene degradation or protein translation inhibition.Thus,as a biological marker for certain diseases,miRNA shows a high degree of specificity.In the present,there are also some studies on miRNA and tuberculosis.Studies have found that in the CD4+T cells of tuberculosis patients,82 miRNAs were up-regulated and 53 miRNAs were down-regulated in active tuberculosis compared with healthy people.In latent tuberculosis patients,33 miRNA were up-regulated and 46 miRNAs were down-regulated.In contrast to patients with latent tuberculosis and active tuberculosis,62 miRNAs were up-regulated and 62 miRNAs were down-regulated.Comparison of the three groups,there were 27 miRNAs have different expression,and it was found that the down-regulated miRNAs in active pulmonary tuberculosis and latent tuberculosis patients have similar expression,indicating that some miRNA are related to the occurrence of tuberculosis infection.It has also been found that some mRNAs can specifically express and play important regulatory roles in Mycobacterium tuberculosis and host cells,such as miR-223,miR-144,and miR-421.At the same time,there are also some studies on miRNA and diatebes.It was find that some microRNAs were related to the insulin expression,such as miR-30d,miR-375,miR-124a2;And some microRNAs affect the insulin secretion,such as miR-375,miR-9,miR-124a2,miR-144,et al?This study focuses on miR-144.MiR-144 has been studied in many other fields,such as miRNA-144,which is an important regulator of erythroid differentiation.It is involved in many cancers,including colon,pancreatic,throat,malignant mesothelioma,and osteosarcoma.And it is involved in regulation of cases of spinal ataxia and Alzheimer's disease.Some studies have found that it is one of the genes that promote cardiomyocyte apoptosis.In addition,there are many studied focused on the respiratory system disease research,studies have shown that miRNA-144 is involved in the regulation of lung cancer,respiratory bronchiolitis,viral pneumonia,asthma,chronic obstructive pulmonary disease,pulmonary fibrosis,pulmonary nodules,pulmonary tuberculosis and other respiratory diseases.In the study between tuberculosis infection and miR-144,miR-144 was find mainly expressed in T cells,and involved in the management of T cells immune function.It can prevent the production of two main immune factors-? interferon and tumor necrosis factor ?.Thus,miR-144 can prevent the proliferation of T cells,and play an important role in the system of tuberculosis infection.Studies have shown that miR-144 is up-regulated in tuberculosis infected patients compared with healthy subjects.The study suggests that miRNA may be involved in the process of tuberculosis infection.In the study between diabetes and miR-144,we find Microrna-144 affects both the expression of insulin genes and the target receptor of insulin.It is highly expressed in peripheral blood of diabetic patients.It mainly regulates the expression of IRS 1 by regulating the control factor of insulin receptor substrate protein 1,which leads to the decrease of insulin function and insulin resistance.Based on the above considerations,we will do this study to further validate the application value of miR-144 in the diagnosis and efficacy evaluation of tuberculosis.Objective1.To detect expression levels of sputum and serum microRNA-144(miR-144)before and after treatment in patients with pulmonary tuberculosis(PTB).2.To measure the levels of miR-144 in sputum and serum of tubercular patients with diabetes and without diabetes.3.To measure the levels of miR-144 and IFN-? in serum and analyze the correlation between them.Method124 cases of PTB patients were collected from our hospital between April 2014 and April 2015.Fifty-three of these patients had sputum positive for bacteria and a cavity on imaging(group A),20 patients had sputum negative for bacteria and a cavity on imaging(group B),and 51 patients had sputum negative for bacteria and no cavity on imaging(group C).Among them,there were 53 tubercular cases complicated with diabetes,including 30 males and 23 females,ranging in age from 20 to 43,with an average age of 28.47±6.98.There were 71 tubercular cases without diabetes,including 40 males and 31 females,ranging in age from 12 to 59,with an average age of 26.40±10.13.There was no significant difference in age and gender between the two groups(P>0.05).One hundred seventeen healthy peoples who attended the hospital for a physical examination were recruited as controls.Quantitative real-time PCR(qRT-PCR)was used to measure the levels of sputum and serum miR-144 before anti-TB treatment and at 1 month after treatment.The statistical data was analyzed using the SPSS 19.0 software.The quantitative data was presented as mean ± standard(mean ± SD).Comparisons of sputum and serum miR-144 expressions before the anti-TB treatment and a month after the treatment were conducted by paired-sample t test.Comparisons of other groups were conducted by the use of independent-samples t test.I2 test was applied to compare count data among groups.Receiver operating characteristic(ROC)curves were employed to evaluate the diagnostic efficiency.Under the ROC curve(AUC)was calculated to assess the diagnostic efficiency of sputum and miR-144.Results1.The expressions of sputum and serum miR-144 in PTB patients before treatmentRT-PCR results showed that before treatment sputum and serum miR-144 expression levels in PTB patients were significantly higher than those of control groups(both P<0.001).Furthermore,before treatment,the expression levels of sputum and serum miR-144 in patients with positive bacteria sputum and cavity PTB were markedly higher than those of patients with negative bacteria sputum and PTB patients without cavity(all P<0.05).2.The diagnostic value of sputum and serum miR-144 for PTBAnalysis of ROC of miR-144 in sputum indicated that AUC was 0.954(95%CI =0.925-0.983,both P<0.001).When cut-off value was 1.185,both the diagnostic sensitivity(86.29%)and specificity(99.15%)reached the highest point.Analysis of ROC of miR-144 in serum presented that AUC was 0.997(95%CI = 0.993-1.000,both P<0.001).When cut-off value was 0.848,both the diagnostic sensitivity(99.19%)and specificity(94.87%)arrived at the highest point.3.The expressions of sputum and serum miR-144 in PTB patients before and after the treatmentAfter the treatment,sputum and serum miR-144 levels in PTB patients significantly decreased compared to those before treatment(both P<0.05).Moreover,compared with control group,the sputum and serum miR-144 levels in PTB patients after the treatment was statistically significant(both P<0.001).The expression levels of sputum and serum miR-144 in patients with positive bacteria sputum and cavity PTB obviously declined after the treatment(all P<0.001).However,for the patients with negative bacteria sputum and cavity PTB,no significant difference of sputum and serum miR-144 expression levels were shown before and after the treatment(all P>0.05).Compared with control groups,there existed statistically significant difference among patients with positive bacteria sputum,negative bacteria sputum,cavity PTB and patients without cavity after treatment(all P<0.05).The sputum and serum miR-144 expression levels in patients with positive bacteria sputum and cavity PTB were not markedly different from those in patients with negative bacteria sputum and cavity PTB(all P>0.05).4.The sputum and serum miR-144 expression levels in patients with different curative effects before and after one month treatmentThe expression levels of sputum and serum miR-144 in those improved PTB patients significantly decreased after the treatment(both P<0.001).The expression levels of sputum and serum miR-144 in those no-improved PTB patients obviously declined after the treatment(both P<0.001).It showed the significant difference of sputum and serum miR-144 expression levels between control groups and improved patients or no-improved patients after the treatment(all P<0.05).Among improved PTB patients,sputum and serum miR-144 levels with positive and negative sputum bacteria and those patients with cavity or not declined significantly after treatment compared to those before treatment.Whereas,the levels among the patients without improvement upregulated significantly before and after the treatment(both P<0.001).The degree of changes of expression levels of sputum and serum miR-144 in improved PTB patients with positive sputum bacteria and cavity PTB were more evident than those of improved PTB patients with negative sputum bacteria and improved PTB patients without cavity with statistical significance(all P<0.05).However,there existed no obvious difference of the expression levels of sputum and serum miR-144 between no-improved patients with positive sputum bacteria and cavity PTB and no-improved patients with negative sputum bacteria and patients without cavity(all P>0.05)5.MiR-144 in the sputum of tubercular patients with diabetic was significantly higher than that of the tubercular patients without diabetic whether before or after treatment.(all P<0.01).6.In the diabetic group,the therapeutic effect between the two groups was compared as follow:(1)Comparison of therapeutic effect between two sputum groups:There was significant difference between group A and group B(p<0.01),while no statistical difference between group B and group C(p=0.069).(2)Comparison of therapeutic effects between two serum groups:There was significant difference between group A and group B(p<0.05),while no statistical difference between group B and group C(p=0.098).7.The difference of IFN-? between the healthy control group and the experimental group was significant before and after one month treatment(all P<0.05).8.MiR-144 was negatively correlated with IFN-? before and after treatment in the experimental group(all P<0.05).Conclusion:1.the results of our ROC indicated that miR-144 in both sputum and serum has high sensitivity and specificity in the diagnosis of tuberculosis,which indicates that the miR-144 can be used as a diagnostic marker for tuberculosis.2.The levels of miR-144 in the sputum and serum of patients with tuberculosis were significantly higher than those in healthy controls,but decreased significantly after anti-tuberculosis treatment.3.Before the anti-tuberculosis treatment,the miR-144 in sputum and serum of B and C group were significantly lower than that in the A group,and C group was significantly lower than the B group.It shows that miR-144 can judge the severity of tuberculosis.4.After treatment,the data showed that the miR-144 in both sputum and serum in each group was lower than that before treatment,which indicated that miR-144 could be used as a therapeutic criterion for the treatment of tuberculosis.5.In the improved group,the miR-144 in the sputum and serum was decreased obviously,while in the pored group,the miR-144 in the sputum and serum has no significant difference.Once again,miR-144 can be used as a therapeutic criterion for the treatment of tuberculosis.6.In the diabetic pulmonary tuberculosis group,miR-144 in the sputum and serum were higher than that in the diabetic pulmonary tuberculosis-free group,whether before or after treatment.7.In the diabetic group,the decreasing degree of miR-144 in the sputum and serum of the patients in group A with positive cavity smear was lower that in the group B without positive cavity smear.8.MiR-144 is negatively associated with IFN-?,and IFN-? is negatively modulated by miR-144. |
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