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Analysis of clarithromycin resistance and its risk f actors in 254 patients with drug-resistant tuberculosis[Objective]Clarithromycin(Clarithromycin,Clr)was once listed as the fifth group of anti-TB drugs by the World Health Organization(WHO).And it may lead to adverse reactions such as prolonged QT interval and be eliminated.In this regard,experts at home and abroad have disputes.This study analyzed 254 cases of drug-resistant tuberculosis patients with clarithromycin resistance and related influencing factors to provide a basis for its clinical application or elimination[Methods]The clinical data and drug sensitivity test results(referred to as drug sensitivity tests")of 254 patients diagnosed with drug-resistant tuberculosis in Beijing Chest Hospital affiliated to Capital Medical University from January 2017 to January 2018 were collected for retrospective analysis.Among them,the sensitivity of 16 kinds of antituberculosis drugs such as clarithromycin was tested using the proportional method.Logistic regression was used to analyze clinically relevant influencing factors for clarithromycin resistance[Results](1)Among 254 patients with drug-resistant tuberculosis,17 multidrug-resistant/extremely multi drug-resistant patients were resistanct to clarithromycin,and the drug resistance rate was 6.69%(17/254),which was lower than the remaining 15 anti-tuberculosis drugs[12.60%(32/254)to 95.67%(243/254)].(2)The drug resistance rates of moxifloxacin(47.06%,8/17),clofazimine(70.59%,12/17),ethambutol(82.35%,14/17),amikacin(52.94%,9/17),p-aminosalicylic acid(76.47%,13/17),p-aminosalicylic acid(88.24%,15/17),and capreomycin(76.47%,13/17)]were higher in the clarithromycin-resistant group than in the sensitive group[moxifloxacin(11.32%,24/212),clofazimine(9.91%,21/212),ethambutol(41.51%,88/212),amikacin(21.70%,46/212),PAS(35.85%,76/212),Pa(52.83%,112/212),capreomycin(28.77%,61/212)],the differences are statistically significant(x2 values were separately 16.721?46.987?10.628?6.793?10.930?7.986?16.370,P values were all<0.05)].(3)Logistic regression result showed that the number of resistant drugs exceeding seven[P=0.004,OR(95%CI)=9.328(2.058?42.290)]was the risk factor for clarithromycin resistance[Conclusion]The resistance rate of clarithromycin in patients with drug-resistant tuberculosis is lower than that of other commonly used anti-tuberculosis drugs,which mostly occurs in patients with multi-drug resistance/broad drug resistance Risk factors.It is suggested that Clr may be used as a drug candidate when Clr-sensitive drug-resistant tuberculosis patients do not have a sufficient number of anti-TB drugs to form a chemotherapy regimenPart ? In vitro activity of clarithromycin on multi-drug r esistant Mycobacterium tuberculosis[Objective]Clarithromycin(Clarithromycin,Clr)belongs to semi-synthetic 14-membered ring second-generation macrolide antibiotics,which can achieve antibacterial effect by inhibiting bacterial protein synthesis.Its minimum inhibitory concentration(MIC)against the standard strain H37Rv of Mycobacterium tuberculosis was 8 ?g/mL,but the activity of Clr on Mycobacterium tuberculosis resistant isolates in vitro was rarely reported.This study determined the in vitro antibacterial activity of Clr and seven other commonly used anti-tuberculosis drugs against Mycobacterium tuberculosis standard strain H37Rv and clinical isolates of MDR tuberculosis,and evaluated the interaction between Clr and five other anti-tuberculosis drugs.Provide a basis for its reasonable choice or elimination[Methods]The standard strain H37Rv of Mycobacterium tuberculosis and three clinical isolates of MDR tuberculosis(MDR-1,MDR-2,MDR-3)were selected and cultured in 7H9 liquid medium.The MABA method was used to determine the minimum bacteriostasis of Clr and seven other anti-tuberculosis drugs(Mfx,Lzd,Cfz,Am,Bdq,EMB,Lfx)against the standard strain H37Rv of Mycobacterium tuberculosis and the clinical isolates of MDR tuberculosis[Results](1)The MIC value of Clr against H37Rv is 6.57 ?g/mL,ranging from 6.57 to 7.07 ?g/mL,which is higher than other seven anti-tuberculosis drugs.The MIC value of Clr against three multidrug-resistant clinical isolates of Mycobacterium tuberculosis(MDR-1,MDR-2,MDR-3)is 2.61 to 7.59 ?g/mL,which is still higher than the other seven anti-tuberculosis drugs;(2)For H37Rv,the combined effect of Clr and Lzd is synergistic(FIC=0.43),the combined effect of Clr and Am is antagonistic(FIC=4.26),and the combined effect of Clr and Mfx,Cfz,Bdq respectively Not relevant(FIC is 1.2,2.26,1.26 respectively).For multidrug-resistant clinical isolates of Mycobacterium tuberculosis(MDR-1),the combined effects of Clr and Lzd,Mfx,and Bdq are additive(FIC is 0.76,0.92,and 0.91,respectively),while Clr is combined with Cfz and Am,respectively The effect is irrelevant(FIC is 1.47 and 1.23 respectively)[Conclusion]The in vitro activity of Clr against H37Rv and clinical isolates of MDR tuberculosis is lower than that of the other seven anti-tuberculosis drugs.The combined application of Clr and Lzd has a synergistic effect on the standard strain of M.tuberculosis H37Rv,and the combined application with Am has an antagonistic effect.For clinical isolates of MDR-TB,Clr combined with Mfx,Lzd and Bdq respectively has an additive effect.It is suggested that Clr can be used as an alternative drug when it is difficult to form MDR-TB,especially XDR-TB chemotherapy regimen,which can facilitate the reduction of Lzd dose,thereby reducing the incidence of adverse reactions and reducing the economic burden of patientsPart? Clarithromycin and clarithromycin-containing chem otherapy regimen on MDR-TB mouse model[Objective]The treatment outcome of MDR-TB is unsatisfactory.According to the 2019 WHO report:The global MDR-TB treatment success rate is 56%,and China's 52%.This may be related to the lack of new anti-tuberculosis drugs,the lack of a reasonable and effective chemotherapy regimen,long course of treatment,large side effects,low patient compliance,and heavy economic burden.During the "Eleventh Five-Year" and "Twelfth Five-Year" period,relying on major national science and technology projects,the treatment of MDR-TB patients was treated with a chemotherapy regimen containing Clr,six drugs,and a course of 18 months.,Higher than the global and China average;and the case fatality rate is 2.33%,lower than the global average.In addition,there are reports in the literature that macrolides,including Clr,have immunomodulatory effects,can suppress the "cytokine storm"caused by pathogen infection,and benefit patients with respiratory diseases through immunomodulatory mechanisms.To this end,this study explored the anti-tuberculosis activity and immune activity of clarithromycin in MDR-TB mouse model,observed and evaluated the efficacy of Clr-containing chemotherapy regimen in MDR-TB mouse model and the dynamic changes of related cytokines during the course of treatment.Provide a basis for Clr's reasonable choice or elimination[Methods]MDR-TB mouse model was established by infecting 157 BALB/c male mice with multi-drug resistant Mycobacterium tuberculosis clinical isolate(MDR-1)bacterial suspension(5 × 106 CFU/ml)and aerosol infection device MDR-TB mice were randomly divided into six groups.Among them,28 in group A(blank control group(CMC group)),20 in group B(Clr treatment group),35 in group C(clr chemotherapy regimen group:Clr-Mfx-Z-Am-EMB-Pto),35 in group D(control group:Mfx-Z-Am-EMB-Pto),35 in group E(control group Lfx-Z-Am-EMB-Pto),and 4 in group F(healthy group).On the 14th day of modeling,MDR-TB mice in group A were given 0.5%sodium carboxymethyl cellulose,MDR-TB mice in group B were given Clr(200 mg/kg),and MDR-TB mice in groups C,D and E Treatment with the above regimen.During the course of treatment,5 mice were sacrificed in each group at the end of each month.MDR-TB mice lungs,spleen gross specimens,quality indicators,bacteriological indicators and other comprehensive evaluation of eff-icacy.MDR-TB mice in groups C,D,and E were discontinued for 3 months after the course of treatment to observe the recurrence of bacteriology.During the course of treatment,at the end of each month,three MDR-TB mice were sacrificed in the B,C,D,and E groups for achieving blood sampling.Using Luminex multi-factor kit to measure the concentration of cytokines:IL-2,IL-4,IL-10,IL-13,IFN-?.[Results](1)20 blank control group MDR-TB mice(CMC group)began to die naturally on the 23rd day after modeling,and a total of 14 died by 30 days after modeling,with a mortality rate of 70%(14/20)and survived The rate is 30%(6/20).Group B MDR-TB mice(Clr group)began to receive Clr treatment on the 14th day of modeling.By the end of four months of treatment,there was no natural death and the survival rate was 100%.Compared with the blank control group MDR-TB mice on the 3rd and 14th day of modeling,? naturally dead MDR-TB mice have more severe pulmonary congestion and edema,and there are densely distributed and unequal gray tuberculosis Focus,some lesions seem to have fusion phenomenon.And the spleen congestion is more serious;?The lung and spleen bacterial load are significantly increased(9.31±0.18,6.03±0.27)(P<0.01);?The body weight(14.69±1.15)is reduced,the lung weight(0.78±0.07),lung The index(5.29±0.43)and spleen index(0.71±0.17)increased(P<0.01),but there was no change in spleen weight(P>0.01).(2)At the end of one month of Clr treatment,compared with the MDR-TB mice in the blank control group,?The lung nodule lesions of the MDR-TB mice in the Clr group were significantly reduced.?The weight of MDR-TB mice in the Clr group increased by 5.69g,and the lung weight,lung index,and spleen index decreased by an average of 0.149g,0.012,and 0.006(P<0.05),but there was no statistical difference in spleen weight(P>0.05).?Lung live bacteria count decreased by 1.12 LgCFU(P<0.05),and there was no significant change in spleen live bacteria count(P>0.05).During the course of Clr treatment,?The lung lesions of MDR-TB mice in the Clr group gradually improved,and a few lesions remained at the end of the 4-month course of treatment,and the spleen did not change significantly.?Body weight,lung weight,spleen weight,lung index,and spleen index were all higher than before treatment(P<0.01).?Lung viable bacteria count decreased month by month,0.98 LgCFU lower than the end of one month of treatment(P<0.05),but the lungs did not reach the sterilized state by the end of 4 months of treatment;there was no significant change in spleen bacterial load.(3)After MDR-TB mice were treated with three chemotherapy regimens,?Compared with the blank control group(CMC group)before treatment(DO)and the end of 1 month of treatment,the size of lung lesions in MDR-TB mice with Clr regimen group It shrinks month by month and the number of lesions also decreases month by month.Although the number of tuberculosis lesions varying from the end to the end of the treatment course was significantly less than that in group E.?Compared with the blank control group(Group A),MDR-TB mice with Clr regimen group(Group C)increased body weight month by month after treatment(P<0.05),lung weight,spleen weight,lung index and spleen index month by month Decrease(P<0.05).However,except for the differences of individual indicators at individual time points,the basic change trend is similar to that of MDR-TB mice in Group D and Group E(P>0.05).? After treatment of MDR-TB mice in the Clr regimen group(group C),the bacterial load of the lungs and spleen at the end of 1 to 4 months decreased month after month(P<0.05)compared with the blank control group(group A)before treatment(DO)and the end of one month of treatment),And its lung bacterial load is lower than Group D and Group E(P<0.05);but its spleen bacterial load is higher than Group D and Group E(P<0.05).Until the end of May,only a few colonies were seen in the lungs,and LgCFU could not be calculated;the spleen load was 0,which reached a sterile state.At the end of June,the lung colony count was 0,reaching a sterile state.However,the lungs in group E did not reach sterility.(4)Three months after the withdrawal of the Clr regimen group(group C),the bacteriological recurrence rate of lung and spleen of MDR-TB mice was 25%(1/4),while the recurrence rate of lung and spleen of group D were both 60%(3/5),the lung bacterial load of group E reached 6.04LgCFU,and the recurrence rate of spleen was 100%(5/5).(5)On the 3rd day of modeling(D-ii)of MDR-TB mice,their serum IL-2 and IL-10 were higher than those in healthy mice,and IFN-y and IL-4 were lower than those in healthy mice(P<0.05),IL-13 is close to healthy mice(P>0.05).After MDR-TB mice were treated with Clr(Group B)and chemotherapy regimen containing Clr(Group C),serum IL-2,IL-4,IL-13 had no significant changes compared with before treatment;IFN-y levels were in Clr After treatment,it gradually increased and reached the end of the treatment period,which was similar to that of healthy mice(P>0.05),but there was no change after treatment with Clr-containing chemotherapy;IL-10 level was slightly lower than before treatment Except for the data of individual indicators at individual time points,this trend is basically similar to Group D and Group E[Conclusion]Clr has a certain anti-tuberculosis activity in vivo,can inhibit t he growth and reproduction of MDR-TB in MDR-TB mice,reduce the bacter ial load of its lungs,promote the absorption and repair of lung lesions,and r elieve its respiratory insufficiency To protect it from death.The chemotherapy regimen containing Clr can quickly reduce the bacterial load of the lungs an d spleen of MDR-TB mice,make it reach a sterilized state at the end of 5-6 months of treatment,promote the improvement of the lesions of the affected organs,and improve the Nutritional status,recurrence rate is lower than the control group.The efficacy is better than the two control schemes.Clr and C lr-containing chemotherapy regimens mainly affect the levels of serum IFN-y and IL-10 in MDR-TB mice.By up-regulating IFN-y levels and down-regulati ng IL-10 levels,the Thl-based immune response is promoted to promote tube rculosis The removal of mycobacteria and the formation of granuloma enable the control of tuberculosis lesions. |
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