理學生理學PPT課件

1、Contents Background Materials and Methods Hypothesis Results Discussion第1頁/共30頁Background Cl- channel c-AMP-PKA CFTRinh-172第2頁/共30頁 CF disease not only a CFTR problem, but also with hyper-absorb Na+. The study of Welsh, denied this view. Welsh used short-circuit current method to do this article.第3頁

2、/共30頁短路電流裝置短路電流 (ISC) (ISC) 定義為穿過上皮并使得跨上皮電壓降為零的電流。鉗制模式1. 電流鉗模式跨上皮電流被鉗制在0A，測量細胞兩側(cè)的電勢差。 2. 電壓鉗模式組織的跨上皮電壓被設(shè)定在一個特定的值，然后接通電壓鉗，跨上皮電流將此電壓維持在預(yù)設(shè)值上。這種模式用于測定由上皮主動轉(zhuǎn)運的離子。第4頁/共30頁第5頁/共30頁短路電流的實驗方法研究上皮正?；虿±砬闆r下的離子轉(zhuǎn)運 研究藥物樣品 液對上皮細 胞Isc的影響阻斷劑法 離子替代法打孔法 第6頁/共30頁阻斷劑法 在加藥反應(yīng)之前加入某種通道的阻斷劑（如Cl-通道阻斷劑DPC等）對組織進行一段時間的孵育，然后再加藥，或在

3、加藥反應(yīng)到高峰的時候再加阻斷劑，看此時藥物的反應(yīng)是否會被抑制。 第7頁/共30頁離子替代法 把正常K-H溶液中的某種離子替換掉（如Cl-用葡萄糖酸根替代），看此時加藥后的反應(yīng)比起在正常K-H中的反應(yīng)是否有變化。第8頁/共30頁打孔法 在細胞兩側(cè)先加入某種離子濃度不相同的溶液（如做K+導就在頂膜側(cè)加入高K+，基膜面加入低K+溶液），然后有目的性地在細胞的某側(cè)加入打孔劑（如制霉毒素，兩性霉素等），將該側(cè)打穿，使該側(cè)外界與細胞內(nèi)相通（即細胞內(nèi)液與該側(cè)外溶液相同），待其穩(wěn)定后，再加入藥物，看此時是否能引起進一步的反應(yīng)。 第9頁/共30頁上皮細胞上皮細胞 生物體內(nèi)依靠不同組織系統(tǒng)的作用有選擇性地將必須的

4、非電解質(zhì)和電解質(zhì)從外環(huán)境吸收或者從內(nèi)環(huán)境分泌出去，所有這些組織系統(tǒng)由大量形態(tài)較規(guī)則，排列緊密的細胞組成，這些多面體細胞組成的幾乎沒有細胞間質(zhì)的薄層稱之為上皮。具有極性：頂膜面和基底膜面 特點具有緊密連接，在形態(tài)上彼此緊密的相連續(xù)不具有毛細血管，依賴附近的結(jié)締組織供給營養(yǎng)和氧氣 主要功能：屏障功能和有方向性物質(zhì)轉(zhuǎn)運功能。第10頁/共30頁Cl- 分泌分泌NKCC cotransporterCFTRNa+ channel基底膜面頂膜面第11頁/共30頁 New born CFTR/ and CFTRF508/F508 pigs increased nasal Vt and Vtamil like

5、persons who have CF. But have no more Na+ absorption than controls. In CF, elevations in basal Vt, basal Isc, Vtamil, and Iscamil are caused by the lack of apical Cl conductance, not by increased Na+ conductance. Measured Gt, Isc, and Vt.第12頁/共30頁Hypothesis How lack of CFTR changes Electrolyte trans

6、port in primary cultures of differentiated human Airway epithelia? Decreased Cl conductance, increased Na+ conductance, or a combination of the two?第13頁/共30頁Materials and Methods Primary Cultures of Human Airway Epithelia. In vitro Human Airway Epithelia. Electrophysiological Measurements.第14頁/共30頁R

7、esults The effects of these compounds on CF and Non-CF Epithelia: 1、 Observations about Gt: Fig.2.第15頁/共30頁Na+ channel activity increased in CF epithelia ?Amiloride sensitive current in epithelia第16頁/共30頁Na+ absorption in CF not exceed that in non-CF.Open-circuit Na+ unidirectional and net Na+ flux

8、ratesFig.3.第17頁/共30頁2、 Observations about IscFig.1.Fig.2.第18頁/共30頁3、 Observations about VtFig.2.第19頁/共30頁Conclusion: Amiloride causes greater Vt and Isc changes in CF Epithelia第20頁/共30頁 Excised Non-CF Epithelia Show Properties Similar to Those in Cultured Epithelia. Fig.2.第21頁/共30頁 Paradoxical Findi

9、ng: Vtamil and Iscamil Not Na+ Conductance Increased in CF. Conclusion: CF/non-CF differences in Basal Vt, Vtamil, and Iscamil ? 第22頁/共30頁Fig.2.Fig.4.Values of basal Vt, Vtamil,and Iscamil did not exceed or were only slightly greater than those in non-CF第23頁/共30頁Fig.5.In Cl-free solutions,basal Vt,

10、Vtamil,and Iscamil did not differ between CF and non-CF.Clfree solutions reduced basal Isc in both non-CF and CF第24頁/共30頁Effects of CFTRinh-172Fig.6.第25頁/共30頁Gt :Same in non-CF and CF Fig.7.第26頁/共30頁Discussion CF Airway Epithelia Have Reduced Anion Conductance but Not Increased Na+ Conductance. The

11、data allow four main conclusions about electrolyte transport in CF vs. non-CF airway epithelia.第27頁/共30頁 First, CF airway epithelia have a markedly reduced trans-epithelial Cl conductance under basal conditions. Second, non-CF epithelia had some constitutive CFTR Cl Conductance under basal conditions. Third, the measurements of Gt, apical Na+ current, and 22Na+ fluxes indicate that Na+ conductance in CF epithelia did not exceed that i