CAJ | 학술논문

目的制备一种人发角蛋白-胶原海绵的三维立体多孔状结构材料,探讨将其作为真皮替代物的可行性.方法将人发在体内具有慢(Z)、中(B)、快(F)三种吸收速度的HHK组分材料编织成网孔为1 mm×1 mm的网格,以酸溶法从牛跟腱中抽提胶原原液,用离心、盐析、透析等方法制备Ⅰ型胶原蛋白溶液,向溶液中滴加胶原总量8%的6-硫酸软骨素(6-GAG)进行初交联.制备好的Ⅰ型胶原溶液与HHK网格混合后放入模具,真空冷冻干燥成海绵状膜,于0.25%戊二醛溶液中浸泡再交联.将制备好的HHK-胶原海绵膜切块(1 cm×1 cm)移植于21只大鼠的真皮与皮下组织之间(实验组),并与单纯胶原海绵膜组(阴性对照)、单纯全层皮肤切开组(空白对照)作对照.于术后3 d和1、2、4、6、8、12周7个不同时间点取材料及其周围组织,行石蜡切片的HE染色、醛复红弹性纤维染色观察及扫描电镜观察检测其组织相容性、血管化能力及材料的体内降解情况.结果人发角蛋白呈棕黄色细丝状.胶原海绵膜为微黄色,半透明状,孔径为50至300μm的三维立体状结构.皮下移植后3 d,各组均表现为中度炎症反应,以中性粒细胞和单核细胞为主.术后1周,炎症反应稍减轻,材料周围可见少量成纤维细胞及新生血管,实验组较对照组明显.术后2周,实验组及阴性对照组,炎性细胞中巨噬细胞增多,实验组成纤维细胞增生活跃,分泌少量胶原物,材料周围新生血管数量增多.HHK及胶原海绵开始降解,创口表皮细胞增殖移行.电镜下可见实验组材料周围包绕的胶原纤维和弹性纤维,人发角蛋白毛小皮降解、剥离;术后4周,成纤维细胞分泌大量粗大的胶原物.创口的表皮细胞增殖移行明显,实验组基本愈合.两种材料崩解明显,胶原海绵网孔内可见巨噬细胞侵入及少量小血管长入.醛复红弹性纤维染色可见短棒或细长条状弹性纤维;术后6至8周,成纤维细胞数量相对减少,分泌胶原物融合成均质粗大,实验组其排列趋于整齐,表皮层愈合良好.人发角蛋白材料中部出现降解裂隙,胶原海绵基本降解完全.实验组醛复红弹性纤维染色可见细长条状弹性纤维;术后12周,人发角蛋白材料降解完全.结论人发角蛋白-胶原海绵膜是物理及生物学性能良好的材料,其组织相容性好,血管化能力强,能刺激细胞增殖,促进受损皮肤愈合,可作为真皮替代物. 目的製備一種人髮角蛋白-膠原海綿的三維立體多孔狀結構材料,探討將其作為真皮替代物的可行性.方法將人髮在體內具有慢(Z)、中(B)、快(F)三種吸收速度的HHK組分材料編織成網孔為1 mm×1 mm的網格,以痠溶法從牛跟腱中抽提膠原原液,用離心、鹽析、透析等方法製備Ⅰ型膠原蛋白溶液,嚮溶液中滴加膠原總量8%的6-硫痠軟骨素(6-GAG)進行初交聯.製備好的Ⅰ型膠原溶液與HHK網格混閤後放入模具,真空冷凍榦燥成海綿狀膜,于0.25%戊二醛溶液中浸泡再交聯.將製備好的HHK-膠原海綿膜切塊(1 cm×1 cm)移植于21隻大鼠的真皮與皮下組織之間(實驗組),併與單純膠原海綿膜組(陰性對照)、單純全層皮膚切開組(空白對照)作對照.于術後3 d和1、2、4、6、8、12週7箇不同時間點取材料及其週圍組織,行石蠟切片的HE染色、醛複紅彈性纖維染色觀察及掃描電鏡觀察檢測其組織相容性、血管化能力及材料的體內降解情況.結果人髮角蛋白呈棕黃色細絲狀.膠原海綿膜為微黃色,半透明狀,孔徑為50至300μm的三維立體狀結構.皮下移植後3 d,各組均錶現為中度炎癥反應,以中性粒細胞和單覈細胞為主.術後1週,炎癥反應稍減輕,材料週圍可見少量成纖維細胞及新生血管,實驗組較對照組明顯.術後2週,實驗組及陰性對照組,炎性細胞中巨噬細胞增多,實驗組成纖維細胞增生活躍,分泌少量膠原物,材料週圍新生血管數量增多.HHK及膠原海綿開始降解,創口錶皮細胞增殖移行.電鏡下可見實驗組材料週圍包繞的膠原纖維和彈性纖維,人髮角蛋白毛小皮降解、剝離;術後4週,成纖維細胞分泌大量粗大的膠原物.創口的錶皮細胞增殖移行明顯,實驗組基本愈閤.兩種材料崩解明顯,膠原海綿網孔內可見巨噬細胞侵入及少量小血管長入.醛複紅彈性纖維染色可見短棒或細長條狀彈性纖維;術後6至8週,成纖維細胞數量相對減少,分泌膠原物融閤成均質粗大,實驗組其排列趨于整齊,錶皮層愈閤良好.人髮角蛋白材料中部齣現降解裂隙,膠原海綿基本降解完全.實驗組醛複紅彈性纖維染色可見細長條狀彈性纖維;術後12週,人髮角蛋白材料降解完全.結論人髮角蛋白-膠原海綿膜是物理及生物學性能良好的材料,其組織相容性好,血管化能力彊,能刺激細胞增殖,促進受損皮膚愈閤,可作為真皮替代物.
목적제비일충인발각단백-효원해면적삼유입체다공상결구재료,탐토장기작위진피체대물적가행성.방법장인발재체내구유만(Z)、중(B)、쾌(F)삼충흡수속도적HHK조분재료편직성망공위1 mm×1 mm적망격,이산용법종우근건중추제효원원액,용리심、염석、투석등방법제비Ⅰ형효원단백용액,향용액중적가효원총량8%적6-류산연골소(6-GAG)진행초교련.제비호적Ⅰ형효원용액여HHK망격혼합후방입모구,진공냉동간조성해면상막,우0.25%무이철용액중침포재교련.장제비호적HHK-효원해면막절괴(1 cm×1 cm)이식우21지대서적진피여피하조직지간(실험조),병여단순효원해면막조(음성대조)、단순전층피부절개조(공백대조)작대조.우술후3 d화1、2、4、6、8、12주7개불동시간점취재료급기주위조직,행석사절편적HE염색、철복홍탄성섬유염색관찰급소묘전경관찰검측기조직상용성、혈관화능력급재료적체내강해정황.결과인발각단백정종황색세사상.효원해면막위미황색,반투명상,공경위50지300μm적삼유입체상결구.피하이식후3 d,각조균표현위중도염증반응,이중성립세포화단핵세포위주.술후1주,염증반응초감경,재료주위가견소량성섬유세포급신생혈관,실험조교대조조명현.술후2주,실험조급음성대조조,염성세포중거서세포증다,실험조성섬유세포증생활약,분비소량효원물,재료주위신생혈관수량증다.HHK급효원해면개시강해,창구표피세포증식이행.전경하가견실험조재료주위포요적효원섬유화탄성섬유,인발각단백모소피강해、박리;술후4주,성섬유세포분비대량조대적효원물.창구적표피세포증식이행명현,실험조기본유합.량충재료붕해명현,효원해면망공내가견거서세포침입급소량소혈관장입.철복홍탄성섬유염색가견단봉혹세장조상탄성섬유;술후6지8주,성섬유세포수량상대감소,분비효원물융합성균질조대,실험조기배렬추우정제,표피층유합량호.인발각단백재료중부출현강해렬극,효원해면기본강해완전.실험조철복홍탄성섬유염색가견세장조상탄성섬유;술후12주,인발각단백재료강해완전.결론인발각단백-효원해면막시물리급생물학성능량호적재료,기조직상용성호,혈관화능력강,능자격세포증식,촉진수손피부유합,가작위진피체대물.
Objective To develop a three-dimensional porous film of human hair keratin (HHK)-collagen sponge complex for use as a dermal substitute. Methods The three components F, B, and Z derived from healthy human hair were weaved into a meshwork and integrated with purified soluble type Ⅰ collagen extracted from bovine tendons to prepare a highly porous film with vacuum freeze-drying followed by secondary cross-linking with glutaraldehyde. The film was grafted beneath the dorsal skin in 21 SD rats (experimental group), with simple collagen sponge serving as the negative control. The rats receiving surgical operation but without graft served as the blank control. The graft and its surrounding tissue were harvested on days 3, 7 and at weeks 2, 4, 6, 8, 12 after implantation for evaluation of tissue compatibility, vascularization and degradation. Results The prepared collagen sponge film was semitransparent and porous. Three to 7 days after grafting, inflammatory reaction was relieved gradually, and several fibroblasts and blood vessels were found adherent to the gratts in the experimental groups. At week 4, the wounds healed in the experimental groups, and the fibroblasts were actively secreting collagen and the film degraded obviously with the appearance of elastic fibers. At weeks 6 and 8, new collagen fibers thickened and assumed regular arrangement, and the collagen sponge films disappeared completely. In the control groups, the changes were less obvious and total HHK degradation occurred till week 12. Conclusion The degradable and absorbable HHK-collagen sponge film has relatively satisfactory tissue compatibility and can accelerate wound healing by stimulating cell proliferation and vascularization,showing the potential as an optimal dermal substitute.