c-MYC-mediated TRIB3/P62+ aggresomes accumulation triggers paraptosis upon the combination of everolimus and ginsenoside Rh2

The mammalian target of rapamycin (mTOR) pathway is abnormally activated in lung cancer. However, the anti-lung cancer effect of mTOR inhibitors as monotherapy is modest. Here, we identified that ginsenoside Rh2, an active component of Panax ginseng C. A. Mey., enhanced the anti-cancer effect of the mTOR inhibitor everolimus both in vitro and in vivo. Moreover, ginsenoside Rh2 alleviated the hepatic fat accumulation caused by everolimus in xenograft nude mice models. The combination of everolimus and ginsenoside Rh2 (labeled Eve-Rh2) induced caspase-independent cell death and cytoplasmic vacuolation in lung cancer cells, indicating that Eve-Rh2 prevented tumor progression by triggering paraptosis. Eve-Rh2 up-regulated the expression of c-MYC in cancer cells as well as tumor tissues. The increased c-MYC mediated the accumulation of tribbles homolog 3 (TRIB3)/P62+ aggresomes and consequently triggered paraptosis, bypassing the classical c-MYC/MAX pathway. Our study offers a potential effective and safe strategy for the treatment of lung cancer. Moreover, we have identified a new mechanism of TRIB3/P62+ aggresomes-triggered paraptosis and revealed a unique function of c-MYC.

Influence of compound salvia injection on oxidative stress and complications in patients with gestational hypertension / 中国基层医药

Objective@#To investigate the influence of compound salvia injection on oxidative stress and complications in patients with gestational hypertension.@*Methods@#From August 2015 to Fabuary 2017, 140 patients with gestational hypertension in Binzhou Central Hospital were selected as study objects, and randomly divided into control group and research group according to the digital table, with 70 cases in each group.The control group was treated with routine treatment, the research group was given routine treatment combined with compound salvia injection.After treatment, SOD, MDA, NO, HMGB1, Cys C, hs-CRP, TNF-α, blood pressure and complications between the two groups were compared.@*Results@#After treatment, the levels of NO[(942.58±80.69)mmol/L vs.(734.51±78.63)mmol/L] and SOD[(124.65±23.81)U/mL vs.(90.64±24.13)U/mL] in the research group were higher than those in the control group, and the MAD level[(10.47±3.14)nmol/L vs.(15.82±3.32)nmol/L]in the research group was lower than that in the control group, the difference were statistically significant between the two groups(t=15.451, 8.394, 9.795, all P<0.05). After treatment, the levels of HMGB1[(1.81±0.43)mg/L vs.(3.89±0.65)mg/L], Cys C[(1.04±0.18)mg/L vs.(1.42±0.27)mg/L], TNF-α[(15.09±3.14)ng/L vs.(21.45±4.78)ng/L] and hs-CRP[(3.04±0.81)mg/L vs.(6.32±1.45)mg/L] in the research group were lower than those in the control group, the differences were statistically significant(t=22.329, 9.798, 9.304, 16.523, all P<0.05). After treatment, the systolic blood pressure[(131.25±5.74)mmHg vs.(139.61±5.59)mmHg] and diastolic blood pressure[(85.23±5.07)mmHg vs.(90.43±5.81)mmHg] in the research group were lower than those in the control group(t=8.730, 5.642, all P<0.05). The cesarean section rate(61.43% vs.87.14%) and intrauterine distress rate(1.43% vs.11.43%) in the research group were lower than those in the control group(χ2=12.115, 5.818, all P<0.05).@*Conclusion@#Compound salvia miltiorrhiza injection has significant control effect on oxidative stress and inflammatory reaction in patients with gestational hypertension, and is beneficial to reduce the complications of childbirth.

Influence of compound salvia injection on oxidative stress and complications in patients with gestational hypertension / 中国基层医药

Objective To investigate the influence of compound salvia injection on oxidative stress and complications in patients with gestational hypertension.Methods From August 2015 to Fabuary 2017,140 patients with gestational hypertension in Binzhou Central Hospital were selected as study objects ,and randomly divided into control group and research group according to the digital table ,with 70 cases in each group.The control group was treated with routine treatment ,the research group was given routine treatment combined with compound salvia injec-tion.After treatment,SOD,MDA,NO,HMGB1,Cys C,hs-CRP,TNF-α,blood pressure and complications between the two groups were compared.Results After treatment,the levels of NO[(942.58 ±80.69)mmol／L vs.(734.51 ± 78.63)mmol／L] and SOD[(124.65 ±23.81)U／mL vs.(90.64 ±24.13)U／mL] in the research group were higher than those in the control group ,and the MAD level [(10.47 ±3.14) nmol／L vs.(15.82 ±3.32) nmol／L] in the research group was lower than that in the control group ,the difference were statistically significant between the two groups(t＝15.451,8.394,9.795,all P＜0.05).After treatment,the levels of HMGB1 [(1.81 ±0.43) mg／L vs. (3.89 ±0.65)mg／L],Cys C[(1.04 ±0.18)mg／L vs.(1.42 ±0.27) mg／L],TNF-α[(15.09 ±3.14) ng／L vs. (21.45 ±4.78) ng／L] and hs-CRP[(3.04 ±0.81) mg／L vs.(6.32 ±1.45) mg／L] in the research group were lower than those in the control group ,the differences were statistically significant (t＝22.329,9.798,9.304,16.523, all P＜0.05).After treatment,the systolic blood pressure[(131.25 ±5.74)mmHg vs.(139.61 ±5.59)mmHg] and diastolic blood pressure[(85.23 ±5.07)mmHg vs.(90.43 ±5.81) mmHg] in the research group were lower than those in the control group(t＝8.730,5.642,all P＜0.05).The cesarean section rate (61.43% vs.87.14%) and intrauterine distress rate(1.43%vs.11.43%) in the research group were lower than those in the control group (χ2 ＝12.115,5.818,all P＜0.05).Conclusion Compound salvia miltiorrhiza injection has significant control effect on oxidative stress and inflammatory reaction in patients with gestational hypertension ,and is beneficial to reduce the complications of childbirth.

Effect of all-trans retinoic acid on transforming growth factor-β1/Notch signaling pathway in injured podocytes induced by Adriamycin / 中华实用儿科临床杂志

Objective To investigate the effect of all-trans retinoic acid (ATRA) on transforming growth factor β1 (TGF-β1)/Notch signaling pathway in injured podocytes induced by adriamycin (ADR) in vitro.Methods Podocytes cultured in vitro were randomly divided into normal group,model group,ATRA treatment control group,12-hour ATRA intervention group and 24-hour ATRA intervention group.Morphological changes were observed by using light microscope.The expressions of TGF-β1,podocin,Notch 1,Jagged 1 mRNA were evaluated through real-time polymerase chain reaction (RT-PCR) and the corresponding proteins were detected by using Western blot.Results (1) No obvious changes between normal group and ATRA treatment control group were revealed as the plump podocytes and distinct outline were found in light microscope,while podocytes in model group showed disordered arrangement,fuzzy boundary,atrophy,hypertrophy and increased cellular debris.Of note,the podocytes in 12-hour ATRA intervention group and 24-hour ATRA intervention group almost returned to normal.(2) In contrast with those in model group,the amounts of TGF-β1,Notchl,Jaggedl mRNA levels decreased in 12-hour ATRA intervention group (1.34 ±0.43 vs.4.16 ±0.31,1.67 ±0.2 vs.4.21 ±0.92,2.08 ±0.27 vs.5.14 ±0.63,q =23.83,11.45,19.67,all P ＜0.05) and 24-hour ATRA intervention group (1.22 ± 0.16 vs.4.16 ± 0.31,1.73 ± 0.53 vs.4.21 ± 0.92,2.08 ± 0.29 vs.5.14 ± 0.63,q =24.85,11.18,19.67,all P ＜ 0.05),and the differences were significant;similar trend was detected in the protein levels (1.04 ± 0.03 vs.4.31 ± 0.10,1.06 ± 0.04 vs.4.47 ± 0.24,1.07 ± 0.04 vs.4.20 ± 0.16,1.06 ±0.03 vs.4.31 ±0.10,1.07 ±0.03 vs.4.47 ±0.24,1.09 ±0.03 vs.4.20 ±0.16,q =163.50,69.61,90.36,162.50,69.40,89.78,all P ＜ 0.05),and the differences were significant;whereas the level of podocin mRNA (1.13 ±0.05 vs.0.40 ± 0.06,1.16 ± 0.03 vs.0.40 ± 0.06,q =36.50,38.00,all P ＜ 0.05) and protein (1.01 ± 0.01 vs.0.44 ±0.01,1.02 ±0.01 vs.0.44 ±0.01,q =180.25,183.41,all P ＜0.05) increased,and the differences were sig nificant.(3) The expressions of Notch1,Jagged1 mRNA were positively correlated with TGF-β1 mRNA (r =0.84,1.00,all P ＜ 0.05),but negatively correlated with podocin mRNA (r =-0.95,-0.94,all P ＜ 0.05) in model group.Conclusions ATRA might alleviate podocyte injury through cutting the expressions of TGF-β1,Notch1,Jagged1 and raising the expression of podocin in injured podocytes induced by ADR.

Cryptotanshinone inhibits TNF-α-induced LOX-1 expression by suppressing reactive oxygen species (ROS) formation in endothelial cells

Cryptotanshinone (CPT) is a natural compound isolated from traditional Chinese medicine Salvia miltiorrhiza Bunge. In the present study, the regulatory effect and potential mechanisms of CPT on tumor necrosis factor alpha (TNF-α) induced lectin-like receptor for oxidized low density lipoprotein (LOX-1) were investigated. Human umbilical vein endothelial cells (HUVECs) were cultured and the effect of TNF-α on LOX-1 expression at mRNA and protein levels was determined by Real-time PCR and Western blotting respectively. The formation of intracellular ROS was determined with fluorescence probe CM-DCFH2-DA. The endothelial ox-LDL uptake was evaluated with DiI-ox-LDL. The effect of CPT on LOX-1 expression was also evaluated with SD rats. TNF-α induced LOX-1 expression in a dose- and time-dependent manner in endothelial cells. TNF-α induced ROS formation, phosphorylation of NF-κB p65 and ERK, and LOX-1 expression, which were suppressed by rotenone, DPI, NAC, and CPT. NF-κB inhibitor BAY11-7082 and ERK inhibitor PD98059 inhibited TNF-α-induced LOX-1 expression. CPT and NAC suppressed TNF-α-induced LOX-1 expression and phosphorylation of NF-κB p65 and ERK in rat aorta. These data suggested that TNF-α induced LOX-1 expression via ROS activated NF-κB/ERK pathway, which could be inhibited by CPT. This study provides new insights for the anti-atherosclerotic effect of CPT.

Research integrity management and cognitive development / 中华医学科研管理杂志

This article first elaborates the connotation of research integrity,and analyzes the causes of the problems,then enumerates the typical examples of research misconduct.Based on those analyses,we explored possible solutions to prevent such problems.Relevant departments should consummate relevant law and regulations to improve the system,to avoid the emergence of research integrity problems,guarantee high quality of research.

Isocryptotanshinone Induced Apoptosis and Activated MAPK Signaling in Human Breast Cancer MCF-7 Cells / 한국유방암학회지

PURPOSE: Isocryptotanshinone (ICTS) is a natural bioactive product that is isolated from the roots of the widely used medical herb Salvia miltiorrhiza. However, few reports exist on the mechanisms underlying the therapeutic effects of ICTS. Here, we report that ICTS has anticancer activity and describe the mechanism underlying this effect. METHODS: The antiproliferative effect of ICTS was determined using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and clonogenic assays. The effect of ICTS on the cell cycle was measured using flow cytometry. Apoptosis was determined by Hoechst 33342 staining, DNA fragmentation assays, and Western blotting for apoptotic proteins. Finally, the effect of ICTS on mitogen-activated protein kinases (MAPKs) was determined by Western blotting. RESULTS: ICTS significantly inhibited proliferation of MCF-7 and MDA-MB-231 human breast cancer cells, HepG2 human liver cancer cells, and A549 human lung cancer cells in vitro. Among the tested cell lines, MCF-7 cells showed the highest sensitivity to ICTS. ICTS significantly inhibited colony formation by MCF-7 cells. Furthermore, exposure of MCF-7 cells to ICTS induced cell cycle arrest at the G1 phase and decreased mitochondrial membrane potential. Hoechst 33342 staining and Western blot analysis for apoptotic proteins suggested that ICTS induced apoptosis in MCF-7 cells. In addition, ICTS activated MAPK signaling in MCF-7 cells by inducing time- and concentration-dependent phosphorylation of JNK, ERK, and p38 MAPK. CONCLUSION: Our results suggest that ICTS inhibited MCF-7 cell proliferation by inducing apoptosis and activating MAPK signaling pathways.

Neocryptotanshinone inhibits lipopolysaccharide-induced inflammation in RAW264.7 macrophages by suppression of NF-κB and iNOS signaling pathways

Neocryptotanshinone (NCTS) is a natural product isolated from traditional Chinese herb Salvia miltiorrhiza Bunge. In this study, we investigated its anti-inflammatory effects in lipopolysaccharide (LPS)-stimulated mouse macrophage (RAW264.7) cells. MTT results showed that NCTS partly reversed LPS-induced cytotoxicity. Real-time PCR results showed that NCTS suppressed LPS-induced mRNA expression of inflammatory cytokines, including tumor necrosis factor α (TNFα), interleukin-6 (IL-6) and interleukin-1β (IL-1β). Moreover, NCTS could decrease LPS-induced nitric oxide (NO) production. Western blotting results showed that NCTS could down-regulate LPS-induced expression of inducible nitric oxide synthase (iNOS), p-IκBα, p-IKKβ and p-NF-κB p65 without affecting cyclooxygenase-2 (COX-2). In addition, NCTS inhibited LPS-induced p-NF-κB p65 nuclear translocation. In conclusion, these data demonstrated that NCTS showed anti-inflammatory effect by suppression of NF-κB and iNOS signaling pathways.

Advances in Proliferation Model of Vascular Smooth Muscle Cells and Inhibitory Effects and Mechanisms of Chinese Herbal Medicine / 世界科学技术-中医药现代化

The proliferation of vascular smooth muscle cells is closely related to the pathogenesis of a variety of diseases, such as atherosclerosis, hypertension, diabetes, vascular restenosis, and etc. A proper model for smooth muscle cell proliferation can be served as an important tool for exploring the molecular mechanisms of smooth muscle cell proliferation and the screening of potential inhibitory agents. Various factors can induce the proliferation of smooth muscle cells with different mechanisms and pathological significance. The ox-LDL, AGEs, angiotensin II (AngⅡ), high glucose, tumor necrosis factor (TNF) and etc. have been successfully applied in the establishment of smooth muscle cell proliferation model. This article summarized the most commonly used models for smooth muscle cell proliferation. The inhibitory effects of Chinese herbal medicine, especially the isolated pure compounds were discussed. It was aimed to provide references for the screening of smooth muscle cells based medications as well as studies on related research and development of Chinese herbal medicine.

Effect of education in pregnant women’s school on primipara mode of delivery and puerperal behavior / 现代临床护理

Objective To investigate the effect of education in pregnant women’s school on primipara mode of delivery and puerperal behavior.Methods Two hundred and twenty cases who delivered in our hospital and paid postpartum visits were divided into observation group and control group equally according to whether they had attended the pregnant women’s school.The observation group included those having attended the school and the control those who had not.The two groups were compared in terms of primipara mode of delivery and puerperal behavior.Results The rates of maternal vaginal delivery and cesarean section rate in observation group were 71.8%and 28.2%,both significantly higher than those of the control group(52.7%and 47.3%)(P<0.01).The rate of caesarean section due to social factors in the observation group was significantly lower than that of the control group(P<0.05).Besides,the observation group had higher rates of puerperiumal shampoo, showering, eating vegetables and fruits,brushing,use of air-conditioners than the control group did(all P<0.01).Conclusions Prenatal maternal health education in the pregnant women’s school affects puerperous behavior.It suggests that prenatal parturients actively participate in related education on maternal health care,which can help establish a good pregnancy and puerperium maternal behavior,reduce maternal cesarean section and thus help them live through puerperium.

Research progress on the anti-cancer effect and mechanisms of the natural tanshinones / 中国药理学通报

Tanshinones are a large class of hydrophobic natural products isolated from traditional Chinese herb Salvia miltiorrhiza Bunge and related plants. Tanshinones include tanshinone ⅡA, cryptotanshinone, tanshinone I, dihydrotanshinone, and so on. Both the tanshinones and the salvianolic acids have been identi-fied as the characteristic and main active ingredients of Salvia miltiorrhiza Bunge with cardiovascular protective activities. Ac-cumulated data in recent years have revealed that tanshinones possess remarkable anti-cancer activities both in vivo and in vitro. In this review, we summarize the latest progress of the an-ti-cancer effect and the underlying mechanisms of tanshinones, particularly with emphasis on tanshinone ⅡA, which might pro-vide reference for the further research and development of these compounds.

Impact of fixed-dose combination of germacrone, curdione, and furanodiene on breast cancer cell proliferation

Herb combination has been very popular in traditional medical prescriptions such as Traditional Chinese Medicine [TCM]. Persistent efforts and attempts have been made to dissect the action mode of TCM in recent years, which has provided certain evidence for inter-herbal interactions. However, the interactions among different components in a single herb have been largely neglected. In this experimental study, the interactions among different components of a single herb were explored. The effect of three main sesquiterpenes [germacrone, curdione, furanodiene] isolated from Curcuma WenyujinY.H. Chenet C Ling on MDA-MB-231 and MCF-7 breast cancer cell proliferation alone or in combination with a fixed-dose-combination was investigated. Furanodiene significantly inhibited cancer cell proliferation while germacrone and curdione showed no effect. Germacrone enhanced furanodiene's anti-proliferative effect. Curdione showed no effect on furanodiene's anti-proliferative effect but partly reversed the anti-proliferative effect of germacrone and furanodiene combined. The morphological and mitochondrial membrane potential [delta [PSI]m] changes showed similar results. However, they demonstrated complicated interactions on the expression of apoptotic-related proteins and key signal transduction proteins. Unpredictable and complex interactions among different components in Curcuma WenyujinY.H.Chenet C Ling may exist. The intra-herb interactions should be taken into consideration when attempts are made to interpret the art of TCM formulation or other similar recipes

A survey of statins application and its efficacy in patients with ischemic stroke / 中华老年医学杂志

Objective To investigate the clinical application of statins for secondary prevention and its efficacy in patients with ischemic stroke.Methods A total of 300 patients with ischemic stroke were divided into statins group (n=91) and control group (n=209) according to whether statins was received or not.Serum lipid parameters,liver and kidney function,carotid artery intimamedia thickness (IMT) and plaque area were compared between the two groups.Results In 209 patients not taking statins,68 cases (32.5％) were not advised by clinic doctor; 55 cases (27.8％)were reluctant to take statins in fear of adverse reactions; 77 cases (36.8％) discontinued stains when serum lipids were normal; 3 cases (1.4％) developed skin allergic reaction and the skin rash that disappeared after stains discontinuance; 3 cases (1.4％) had elevated liver enzyme levels and the enzyme levels were declined to normal after stains discontinuance.The levels of total cholesterol (TC),triglyceride (TG) and low density lipoprotein cholesterol (LDL-C) were significantly lower in statin group than in control group [(4.93±0.45) mmol/L vs.(5.87±0.38) mmol/L,(1.68±0.61)mmol/L vs.(2.01±0.58) mmol/L,(2.49±0.57) mmol/L vs.(3.76±0.44) mmol/L,t=18.60,4.46,20.94,P=0.000,0.007,0.000,respectively].High density lipoprotein (HDL-C) level was significantly higher in stain group than in control group [(1.36±0.31) mmol/L vs.(1.03±0.25)mmol/L,t=9.75,P=0.001].There were no significant differences in liver and kidney function parameters such as alanine aminotransferase,aspartate aminotransferase,alkaline phosphatase,serum total bilirubin,direct bilirubin,indirect bilirubin,glutamyl transferase,blood urea nitrogen and serum creatinine between the two groups (all P＞0.05).The carotid IMT and carotid atherosclerotic plaque area were significantly lower in stain group than in control group [(1.62 ± 0.23) mm vs.(1.74±0.31) mm,(0.57±0.12) cm2 vs.(0.65±0.18) cm2,t=3.32,3.88,P=0.008,0.002,respectively].Conclusions Most patients with ischemic stroke are still not taking statin for secondary prevention.Statins can improve the blood lipid levels and carotid arteriosclerosis status,and no obvious adverse reactions are observed.

Inspirations from natural products based drug research and development for Chinese medicine research--analysis of natural products recoded in TTD / 药学学报

Natural product is an important source of new drug research and development (R&D). Traditional Chinese medicine (TCM) innovation is the key step for its modernization and internationalization. However, due to the complexity of TCM, there are many difficulties and confusions in this process. Target-based drug discovery is the mainstream model and method of R&D. TTD, short for therapeutic target database, is developed by National University of Singapore. Besides a large amount of information on drug targets, the database also contains considerable information related to natural products. This paper briefly introduces the TTD, analyzes the natural products derived drugs/compounds recorded in TTD, which we think might provide some inspiration for the innovation of TCM.

Multislice spiral CT and digital subtraction angiography in interventional therapy of hepatocellular carcinoma / 中国基层医药

Objective To investigate the value of multislice spiral CT(MSCT,)in interventional therapy of the hepatocellular carcinoma(HCC)emphasising on transcatheter hepanc arterial chemoembolization(TACE).Methods MSCT were performed in 60 cases of HCC before interventional procedure,CT findings of hepatic artery phase,portal venous phase and hepatic venous phase were observed respectively,among which CTA were done in 15 cases,and the anatomy of celiacartery and its branches were observed in 45 cases.The schemes of interventional therapy were worked out according to the findings of MSCT.Results MSCT showed 250 lesions,10 cases of tumor thrombosis in portal vein and 19 cases of hepatic arterioportal shunt.There was no significant difference between MSCT and digital subtraction angiography(DSA)in positive rate of in showing number of tumor or tumor thrombosis in portal vein(P＞0.05),but the 3D construction of celiac artery branches in CTA was better than that in DSA,while angles between celiac artery and abdominal aorta in MSCT were more convenient than that in DSA.MSCT showed 5 eases of hepatic artery original abnormality,according to that in DSA.Conclusion MSCT is of importance for guidance of interventional therapy of hepatocellular carcinoma.

Catheter-related Sepsis: Clinical and Etiologic Analysis / 中华医院感染学杂志

OBJECTIVE To investigate the clinical and etiologic characteristics and treatment of central venous catheter-related(sepsis).METHODS Forty cases of patients with central venous catheter-related sepsis and their clinical manifestation,etiologic profiles and outcome of treatment were retrospectively analyzed.RESULTS Forty six strains were isolated including 23 strains of Gram-positive cocci,19 of Gram-negative bacilli and 4 of fungi.The most frequent(isolates) were Staphylococcus epidermidis.CONCLUSIONS The most common organism causing CRS is S.epidermidis;the key preventive measure is to avoid inner and outer pollution of catheter;antibiotic lock-technique can be taken for the treatment of uncertain CRS.If it is no effect after 24-48 hours,it is necessary to remove venous catheter promptly.

Sensitivity of Vibrio vulnificus to Common Antimicrobials / 中华医院感染学杂志

OBJECTIVE To determine the sensitivity of Vibrio vulnificus to 23 kinds of antibacterial drugs,and offer the evidence for clinical V.vulnificus infections treatment.METHODS Isolated 11 strains of V.vulnificus from the patients blood and blisters of infected parts,using ATB automatic identification and susceptibility testing to measure the vitro drug sensitivity of the bacteria.RESULTS The susceptibility of the bacteria to the 22 kinds of commonly used antibiotics showed 100% sensitivity,the rate of ciprofloxacin sensitivity was 72%.CONCLUSIONS It is sensitive to commonly used semi-synthetic penicillin,cephalomycin,carbapenem,aminoglycosides,the third generation cephalosporins,fluoroquinolones and so on,all these above drugs may be chosen to treat the V.vulnificus infection.

Systematic pharmacological evaluation of material basis for efficacy of Chinese herbal formula / 中草药

Systematic pharmacological evaluation of material basis for the efficacy of Chinese herbal formula is one of the key issues in Chinese materia medica(CMM),which is in urgent need of solution.However,its scientific connotation is still controversial in academic circles.The material basis for the efficacy of Chinese herbal formula is the total non-endogenous bioactive compound(derived either directly or indirectly from this formula) exerting the therapeutic effects,which is generated by the interaction of the formula with human body and reaches certain concentration in the target organs or tissues.Systematic evaluation of material basis for the efficacy of Chinese herbal formula is the cornerstone of modernization and internationalization of CMM,while the quality control of CMM is its prerequisite.