Cov nyhuv ntawm Cistanche Deserticola Polysaccharide Ntawm Lymphocyte Proliferation

Wang Xiangyan, Qi Yun, Cai Runlan, Li Xiaohong, Yang Meihua, Shi Yue

(Institute of Medicinal Plants, Peking Union Medical College, Suav Academy of Medical Sciences, Beijing 100193)

Lub hom phiajTo kawm cov nyhuv ntawmCistanche deserticola polysaccharide(CDPS) ntawm lymphocyte proliferation hauv nas. 

Cov txheej txheemMTT kev soj ntsuam tau siv los txheeb xyuas qhov kev loj hlob ntawm splenic lymphocytes hauv cov nas. Cyclophosphamide (Cy) tau siv los hloov cov qauv tsiaj ntawm cov tsiaj txhu tiv thaiv kab mob, thiab cov spleen thiab thymus indices ntawm cov tsiaj ib txwm muaj thiab tiv thaiv kab mob. Qhov kev ua ntawm interleukin -2 (IL-2) tau ntsuas los ntawm kev soj ntsuam thymocyte proliferation. Cov txiaj ntsigCistanche deserticola polysaccharidetxhawb nqa kev loj hlob ntawm mitogen (ConA thiab LPS) ua kom cov lymphocytes thiab cov hlwb tsis ua haujlwm, thiab txhawb kev tso tawm ntawm IL-2 hauv lymphocytes. Intraperitoneal tswj tau pom tiasCistanche deserticola polysaccharidence qhov poov xab Performance index ntawm ib txwm thiab immunocompromised nas, thiab kuj muaj ib tug tseem ceeb antagonistic nyhuv ntawm kev txo qis ntawm thymus index tshwm sim los ntawm Cy. XausCistanche deserticola polysaccharidetuaj yeem txhawb kev loj hlob ntawm splenic lymphocytes hauv cov nas, uas yuav cuam tshuam rau nws txoj kev txhawb nqa ntawm IL-2 secretion.

Suab puam nyob cistanche

[Cov lus tseem ceeb]Cistanche deserticola polysaccharide; Pleev index; Thymus index; Kev loj hlob ntawm lymphocytes; Interleukin -2

Nroj tsuag polysaccharides tau nyiam thoob plaws lub ntiaj teb vim lawv cov kab mob immunomodulatory thoob plaws thiab tsis tshua muaj tshuaj lom [1].Cistanche deserticola ma, hu ua "Desert Ginseng", yog scaly, qhuav, fleshy qia ntawm Cistanche deserticola, Ib tsob nroj ntawm Orobaceae, uas muaj cov tshuaj siv tshuaj ntau heev.Nws cov dej decoction tuaj yeem ua rau nce phagocytic feem pua ​​thiab phagocytic Performance index ntawm macrophages.Cov kev tshawb fawb ntxiv tau pom tias nws cov khoom siv hauv paus yog tag nrho polysaccharide ntawm Cistanche deserticola[3]. Zeng Qunli et al. [4] tau pom tias ib feem ntawmCistanche deserticola polysaccharidemuaj qhov cuam tshuam loj heev ntawm nas thymic lymphocytes hauv vitro. Cov nyhuv ntawmCistanche deserticola polysaccharideKev loj hlob ntawm lymphocytes tau pom hauv vivo thiab hauv vitro.

Molecular mis ntawm Cistanche deserticola

1. Cov khoom siv thiab cov txheej txheem

1.1 Tshuaj thiab tshuaj reagent

Cov polysaccharide los ntawm Cistanche deserticola yog muab los ntawm Lub Tsev Haujlwm Saib Xyuas Kev Noj Qab Haus Huv thiab Tshuaj Kho Mob ntawm Lub Tsev Haujlwm Saib Xyuas Tshuaj thiab Tsob Ntoo, Tsev Kawm Ntawv Kho Mob. Nws yog muab rho tawm los ntawmCistanche extract hmoov, thiab cov ntsiab lus polysaccharide yog 87 feem pua.

Cistanche extract hmoov

Ua ntej siv hauv vivo thwmsim, yaj thiab dilute nrog physiological saline mus rau qhov xav tau concentration, thiab boil rau kom tsis muaj menyuam. Ua ntej siv hauv vitro sim, RPMI -1640 kab lis kev cai nruab nrab tau yaj thiab diluted, thiab kev ua kom tsis muaj zog siab tau ua. Lipopolysaccharide (LPS), concanavalin (ConA), dexamethasone (DEX), thiab MTT yog txhua yam khoom ntawm Sigma Corporation. Cyclophosphamide, Sino-Asmeskas kev koom tes ntawm Shanxi Taisheng Pharmaceutical Co., Ltd. Fetal bovine serum, RPMI1640 kab lis kev cai nruab nrab, thiab trypsin yog tag nrho cov khoom Gibco. Lwm cov reagents yog cov tshuaj ntsuam xyuas hauv tsev.

1.2 Tsiaj

BALB c cov nas me (SPF qib) yog muab los ntawm Viton Lihua Experimental Animal Technology Co., Ltd. [SCXK (Beijing) 2002-2003]. Cov tsiaj qib SPF raug tsa hauv chav thiab pub dawb thiab haus dej.

1.3 Apparatus

Multikan Ascent enzyme marker, ib yam khoom ntawm Thermo Electron Company, USA. Leici PHS-3B Precision pH Meter, Shanghai Precision Scientific Instrument Co., Ltd. Napco 5410 carbon dioxide incubator, NAPCO, USA. BCN-1360 Ultra Clean Workbench, Beijing Donglianhar Instrument Manufacturing Co., Ltd. XDS-1B Inverted Microscope, Chongqing Optoelectronic Instrument Co., Ltd.

1.4 Method

1.4.1 Cov nyhuv ntawm lymphocyte proliferation [5]:

Kev npaj cov spleen lymphocytes los ntawm BALL c nas, siv RPMI-1640 los kho cov cell concentration mus rau 1 × 107 mL, inoculated rau ntawm 96 zoo kab lis kev cai phaj, nrog 100 cell suspensions ntxiv rau txhua qhov dej μ 50. Nws yog co -cultured nrog txawv concentrations ntawmCistanche deserticola polysaccharide(qhov kawg concentration ntawmCistanche deserticola polysacchariderau kev loj hlob ntawm nonactivated lymphocytes yog 6.25, 12.5, 25, 50, thiab 100 mg / L nyob rau hauv kev txiav txim; qhov kawg concentration ntawmCistanche deserticola polysacchariderau kev loj hlob ntawm ConA-activated T lymphocytes yog 3.9, 7.8, 15.6, 31.2, thiab 62.5 mg / L nyob rau hauv kev txiav txim; thiab qhov kawg concentration ntawmCistanche deserticola polysacchariderau kev loj hlob ntawm LPS activated B lymphocytes yog 1.56, 3.13, 6.25, 12.5, thiab 25 mg / L nyob rau hauv kev txiav txim) thiab ConA los yog LPS, RPMI -1640 nruab nrab yog siv los tswj. Tag nrho cov ntim ntawm txhua lub qhov yog 200 μ L, thaum ntxiv mitogen ConA thiab LPS (qhov kev tswj yog RPMI-1640 uas muaj cov concentrations ntawm ConA lossis LPS), qhov kawg ntawm ob qho tib si yog 2.5 mg L thiab 10 mg L, raws. Peb qhov sib npaug tau ua rau ntau yam kev kho mob. Muab cov phaj kab lis kev cai tso rau hauv 37 degree, 5 feem pua ​​​​CO2 saturated humidity incubator rau 68 teev, ntxiv MTT, thiab txuas ntxiv mus rau kab lis kev cai rau 4 teev. Ntsuas tus nqi A540 nm nrog tus cim enzyme siv MTT txoj hauv kev txhawm rau kuaj pom kev loj hlob. Thaum sib piv txhua pab pawg, hloov tus nqi A540nm rau hauv lymphocyte proliferation tus nqi raws li cov qauv hauv qab no:lymphocyte proliferation rate= Ib qho piv txwv A tswj × 100 feem pua ​​.

Cov teebmeem ntawm Cistanche - Txhim kho kev tiv thaiv

1.4.2 Qhov cuam tshuam rau qhov hnyav ntawm lub cev tiv thaiv kab mob hauv cov nas ib txwm muaj [6,7]:

Young BALB c nas (txiv neej, 4-ib lub lis piam, nrog lub cev hnyav ntawm 10-15 g)) tau muab faib ua pawg tswj dawb (saline) thiab pawg ntau npaum li cas (125 mg (kg · bw) thiab 250 mg (kg · bw)), nrog rau 11 nas nyob rau hauv txhua pab pawg. Cov tshuaj tau muab tshuaj intraperitoneally ib hnub ib zaug rau 7 hnub sib law liag. Cov nas raug tua tom qab lub ncauj tsev menyuam dislocation. Cov poov xab thiab thymus raug muab tshem tawm, rog thiab fascia cov ntaub so ntswg raug tshem tawm, thiab cov kua saum npoo tau qhuav nrog cov ntawv lim. Tom qab qhov hnyav, tus po index =100 × Spleen hnyav (mg) Lub cev hnyav (mg); Thymus index =100 × Thymus Nyhav (mg) Lub Cev Nyhav (mg) Xam cov spleen index thiab thymus index rau txhua 100 mg ntawm lub cev hnyav, raws li.

1.4.3 Qhov cuam tshuam rau lub cev tiv thaiv kab mob hnyav hauv cov nas uas muaj kab mob tiv thaiv kab mob [7,8]:

Hluas BALB c nas (txiv neej, 6 lub lis piam, lub cev hnyav 15-20 g) tau muab faib ua 12 pawg raws li lub cev hnyav. Tsuas yog rau pawg tswj hwm ib txwm muaj (kev txhaj tshuaj intraperitoneal ntawm qhov sib npaug ntawm lub cev saline), lwm pab pawg tau txhaj tshuaj Cy 50 mg (kg · bw) txhua hnub intraperitoneally, thiab cov tshuaj ntsuam xyuas tau muab tshuaj intraperitoneally [125 thiab 250Mg (kg· bw)], pawg tswj hwm ib txwm muaj thiab pawg qauv tau muab tib lub ntim ntawm physiological saline rau 13 hnub sib law liag, ib hnub ib zaug. Cov kev hloov pauv ntawm cov tsiaj hnyav tau pom thaum lub sijhawm sim. 24 teev tom qab kev tswj hwm zaum kawg, tus po thiab thymus indices tau ntsuas thiab suav siv tib txoj kev raws li hauv 1.4.2.

1.4.4 Kev cuam tshuam rau IL-2 kev ua:

① Induction ntawm IL-2 [9]: Ntxiv 50 rau txhua qhov dej hauv 96-zoo cell phaj μ L Qhov sib txawv ntawmCistanche deserticola polysaccharide(qhov kawg concentrations nyob rau hauv kev txiav txim: 3.91, 7.82, 15.63, 31.25, 62.5, 125 mg / L, ntxiv 50 mg / L rau hauv kev tswj zoo μ L kab lis kev cai nruab nrab) thiab 50 μ L ConA (kawg concentration 5 mg / L), ntxiv 1 ntxiv rau qhov dej × 106 spleen hlwb. Incubate nyob rau hauv ib tug 37 degree, 5 feem pua ​​CO2 saturated humidity incubator rau 24 teev, centrifuge rau 20 feeb ntawm 2500 r min, thiab coj lub cell supernatant thiab khaws cia ntawm - 20 degree rau standby. ② IL-2 kev tshawb nrhiav kev ua haujlwm [9,10]: Cov txheej txheem yooj yim yog ua raws li cov ntaub ntawv [9], thiab kev hloov kho tsim nyog tau ua raws li cov ntaub ntawv [10]. Raws li kev npaj ntawm BALB c nas thymocytes ncua kev kawm, 96 lub cell kab lis kev cai phaj, 1 inoculation ib qhov dej × 106 thymocytes, thaum ntxiv 50 hlwb ib qhov dej μ L ConA (qhov kawg concentration 2.5 mg / L), thiab tom qab ntawd ntxiv cov npaj IL {{34 }} supernatant 50 μ L (ntxiv 50 rau lub qhov khoob tswj qhov μ L kab lis kev cai tov), ​​incubate ua ke rau 54 teev, ntxiv MTT, txuas ntxiv incubating rau 4 teev, centrifuge rau 10 mus rau 15 feeb ntawm 1000 r min, pov tseg cov supernatant, thiab ntxiv DMSO 100 rau ib qhov dej μ L, co nrog micro oscillator rau 8 min, thiab ntsuas tus nqi A540 nm nrog tus cim enzyme.

1.5 Kev Tshawb Fawb Txog Kev Tshawb Fawb

Cov ntaub ntawv tau qhia raws li qhov nruab nrab thiab tus qauv sib txawv (x ± s). Microsoft Excel software tau siv los sau cov ntaub ntawv, thiab kev xeem t-test tau ua. P<0.05 and P<0.01 were significant differences, both of which were statistically significant.

2. Kev tshwm sim

2.1 Cov nyhuv ntawm lymphocyte proliferation

Raws li tuaj yeem pom los ntawm Table 1, ntxivCistanche deserticola polysaccharidemus rau kab lis kev cai nruab nrab ntawm nas spleen lymphocytes kab lis kev cai hauv vitro ntawm qhov siab tshaj 25 mg / L tuaj yeem txhawb nqa lymphocyte proliferation, qhia tiasCistanche deserticola polysaccharidenws tus kheej muaj cov nyhuv mitotic. Tib lub sijhawm,Nws tuaj yeem pom los ntawm Table 2 thiab Table 3 tias CDPS muaj qhov cuam tshuam loj rau kev loj hlob ntawm ConA-induced T lymphocytes lossis LPS-induced B lymphocytes..

Tab.1 Lub zog ntawmCistanche deserticola polysaccharidentawm kev loj hlob ntawm nonactivated lymphocytes (x ±s, n =3)

Nco tseg: piv nrog rau pawg tswj hwm,＊P <0.05. 

Tab.2 Lub zog ntawmCistanche deserticola polysaccharidentawm kev loj hlob ntawm Tlymphocytes induced los ntawm ConA (x ±s, n =3)

Nco tseg: piv nrog ConA pab pawg,＊P <0.05, ＊＊P <0.01.

Tab.3 Lub zog ntawmCistanche deserticola polysaccharidentawm kev loj hlob ntawm Blymphocytes induced los ntawm LPS (x ±s, n =3)

Nco tseg: piv nrog pawg LPS,＊P <0.05 , ＊＊P <0.01. 

2.2 Qhov cuam tshuam rau qhov hnyav ntawm lub cev tiv thaiv kab mob hauv cov nas ib txwm muaj

Raws li tau pom los ntawm Table 4,Cistanche deserticola polysaccharidetuaj yeem nce tus po index ntawm ib txwm BALB c cov nas me tab sis tsis muaj kev cuam tshuam rau lub caj pas thymus.

2.3 Qhov cuam tshuam rau lub cev tiv thaiv kab mob hnyav hauv cov nas uas muaj kab mob tiv thaiv kab mob

Los ntawm Table 5,Nws tuaj yeem pom tau tias tus po thiab thymus organ indexes ntawm nas nyob rau hauv pawg Cy immunosuppressive qis dua cov hauv pawg tswj, thiabCistanche deserticola polysaccharide(125, 250 mg / kg) tuaj yeem txhim kho tus po thiab thymus indexes.

2.4 Qhov cuam tshuam rau IL-2 kev ua haujlwm

Raws li tau pom los ntawm Table 6,thaum koob tshuajCistanche deserticola polysaccharidence mus txog 7.82 mg / L, nws tuaj yeem nce qhov tso tawm ntawm IL -2 los ntawm nas spleen hlwb.

Tab.4 Cov nyhuv ntawmCistanche deserticola polysaccharidentawm qhov hnyav ntawm lub cev tiv thaiv kab mob hauv cov nas ib txwm muaj (x ±s, n =11)

Nco tseg: piv nrog rau pawg tswj hwm,＊＊P <0.01.

Tab.5 Cov nyhuv ntawmCistanche deserticola polysaccharidentawm qhov hnyav ntawm lub cev tiv thaiv kab mob hauv cov nas uas tsis muaj zog tiv thaiv kab mob (x ±s, n =12)

Nco tseg: piv nrog rau pawg tswj hwm,## P <0.01;compared with the model group , ＊＊P <0.01.

Tab.6 Lub zog ntawmCistanche deserticola polysaccharidentawm IL -2 ntau lawm los ntawm splenic lymphocytes hauv nas (x ±s, n =3)

Nco tseg: piv nrog rau pawg tswj hwm,＊＊P <0.01. 

3. Kev sib tham

ConA thiab LPS-induced splenic lymphocyte proliferation nyob rau hauv nas yog ib qho kev sim hauv vitro los kuaj xyuas cov haujlwm ntawm T thiab B lymphocytes thiab yog ib qho tseem ceeb qhia txog kev tiv thaiv kab mob. Kev tshawb fawb tau pom tias thaum kawg concentration ntawmCistanche deserticola polysaccharideyog siab dua 25 mg / L, nws tuaj yeem txhawb nqa lymphocyte proliferation, qhia tias nws muaj qee yam mitotic nyhuv. Nyob rau hauv lub xub ntiag ntawm mitogen (ConA los yog LPS), ib koob me me ntawmCistanche deserticola polysaccharidetuaj yeem txhim kho kev tiv thaiv ntawm lub cev, nthuav tawm cov haujlwm tseem ceeb ntawm kev loj hlob. Peb paub tias ConA thiab LPS feem ntau ua rau kev loj hlob ntawm T lymphocytes lossis B lymphocytes, feem. Cov kev tshawb fawb tau qhiacistanche hmoov cov txiaj ntsig kev noj qab haus huv: Cistanche deserticola polysaccharidetuaj yeem txhawb nqa T lymphocyte proliferation thiab B lymphocyte proliferation, tab sis nws proliferative nyhuv ntawm B lymphocytes ho muaj zog dua li ntawm T lymphocytes.

Cistanche hmoov cov txiaj ntsig kev noj qab haus huv-Improve tiv

Nyem qhov no mus saib Cistanche Improve Immunity khoom

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Lub thymus thiab spleen yog lub cev tiv thaiv kab mob tseem ceeb hauv cov tsiaj. Cov tshuaj tiv thaiv kab mob hauv tus po yog cov kab mob B lymphocytes uas paub tab, thaum lub thymus feem ntau tsis paub tab T lymphocytes. Thymus thiab spleen indices tuaj yeem cuam tshuam txog kev txhim kho ntawm lub cev tiv thaiv kab mob thiab kev ua haujlwm ntawm lub cev tiv thaiv kab mob. Kev txhaj tshuaj Intraperitoneal ntawm Cistanche deserticola polysaccharide tuaj yeem ua rau kom cov poov xab index ntawm cov nas ib txwm muaj thiab tiv thaiv kab mob thiab muaj qhov cuam tshuam tseem ceeb ntawm thymic atrophy induced los ntawm Cy hauv cov nas.Cistanche deserticola polysaccharidetsis muaj qhov cuam tshuam tseem ceeb ntawm thymic Performance index hauv cov tsiaj ib txwm, uas yog raws li cov txiaj ntsig ntawm kev sim hauv vitro uasCistanche deserticola polysaccharidetxhawb kev loj hlob ntawm B lymphocytes es tsis yog T lymphocytes.

Cistanche ntxiv nyob ze kuv—Txhim khotiv thaiv kab mob

IL-2 yog cytokine tsim los ntawm T lymphocytes los ntawm G1 theem ntawm lub voj voog ntawm tes mus rau theem S. Nws yog ib qho tseem ceeb ntawm T cell loj hlob yam uas ua raws li autocrine kev loj hlob yam. Tsis tas li ntawd, IL -2 tuaj yeem txhawb B cell proliferation thiab antibody synthesis [11,12].Cistanche deserticola polysaccharidetxhawb kev tso tawm ntawm IL-2 los ntawm lymphocytes, uas yog pom tseeb hais txogCistanche deserticola polysaccharidetxhawb kev loj hlob ntawm splenic lymphocytes.

Kev tshawb fawb kho mob qhia tau hais tias Cistanche muaj polysaccharides,Echinacosides thiab Acteoside, uas tuaj yeem qhib cov lymphocytes thiab immprove kev tiv thaiv ntawm body; cov khoom siv ua haujlwm ntawm cistanche Phenylethanoid Glycosides muaj cov txiaj ntsig tseem ceeb ntawm kev rov ua haujlwm ntawm tes tom qab 60 Co gamma ray puas, thiab aslo tuaj yeem txhim kho lub cev tsis muaj zog tiv thaiv hluav taws xob puas tsuaj.Noj Cistanche deserticola ntxiv txhua hnub tuaj yeem ua tiav cov txiaj ntsig ntawm kev txhim kho kev tiv thaiv.

Cistanche ntxiv nyob ze kuv—Txhim kho kev tiv thaiv

siv

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