软骨细胞诱导骨髓基质干细胞构建带内支撑的组织工程化软骨

目的探讨以聚羟基乙酸(PGA)包裹特定形态的医用假体材料多孔高密度聚乙烯(HDPE,商品名为MEDPOR)为支架,应用软骨细胞诱导骨髓基质干细胞(BMSCs),共培养构建特定形态的带内支撑组织工程化软骨医用假体的可能性。方法以直径3mm、长5mm的圆柱形HDPE,外裹1mm厚PGA为支架,将体外分别培养的新生猪BMSCs和耳郭软骨细胞按7:3混合,以10×107/ml细胞浓度接种于支架上,同时以相同浓度的单纯软骨细胞和单纯BMSCs分别接种,作为阳性对照组(PC组)和阴性对照组(NC组)。经体外培养2周及在裸鼠皮下移植4、8周后取材,行大体观察、组织学、组织化学及免疫组化检测。结果各组细胞均与材料黏附良好。实验组和阳性对照组均形成了大体形态良好的HDPE-软骨复合体,内支撑的HDPE与外层软骨结合紧密。组织学可见成熟的软骨陷窝结构,软骨渗入HDPE孔隙内部、异染基质及Ⅱ型胶原呈强阳性表达。结论以HDPE为内支撑,外裹PGA的支架,接种混合细胞,可于皮下构建特定形态、组织学良好的HDPE-软骨复合体。     

构建带内支撑组织工程软骨的实验研究

目的探讨以医用多孔高密度聚乙烯(Medpor)为内支撑外裹聚羟基乙酸(PGA)的支架,接种软骨细胞后,于体内构建Medpor软骨复合体的可能性。方法实验组:以直径3mm的Medpor外裹2mmPGA为支架,接种新生猪关节软骨细胞(5×107/ml),经体外培养2周及裸鼠皮下移植6周后取材,行大体、组织学、Ⅱ型胶原免疫组织化学和生物化学检测;PGA对照组:以直径8mm的单一PGA为支架,细胞接种与培养同实验组;单纯支架组:利用实验组支架,不接种细胞行体内移植。结果实验组:形成大体形态良好的Medpor-软骨复合体,内部的Medpor与外层软骨结合紧密,组织学可见成熟软骨陷窝并渗入Medpor孔隙内部、异染基质、Ⅱ型胶原表达阳性;PGA对照组:形成的软骨存在"空心"现象;单纯支架组:无软骨形成。结论利用Medpor与PGA复合的支架,接种软骨细胞后,可于体内构建出特定形状、结构与组织学良好的Medpor软骨复合体,克服了组织工程软骨的"空心"现象。     

以骨髓基质干细胞构建带内支撑的组织工程化软骨

目的 探讨构建特定形态带内支撑组织工程化软骨的医用假体的可能性.方法 以直径3 mm、长5 mm的圆柱状多孔高密度聚乙烯(Medpor)外裹厚1 mm的聚羟基乙酸为支架,将体外培养的骨髓基质干细胞(bone marrow stromal cells,BMSCs).按10×107/ml的细胞浓度均匀接种于支架,常规培养液培养5 d后,用含诱导因子的培养液立体诱导4周,同时以相同浓度的软骨细胞和BMSCs分别接种,常规体外培养4周作为阳性对照组和阴性对照组,分别种植于裸鼠皮下,4、8周后取材,行大体观察、组织学、组织化学、免疫组化及糖氨聚糖(GAG)定量等检测.结果 各组细胞均与材料粘附良好.实验组和阳性对照组均形成了大体形态良好的Medper-软骨复合体,内部的Medper与外层软骨结合紧密.组织学可见成熟软骨陷窝并渗入Medper孔隙内部、异染基质及Ⅱ型胶原表达,实验组GAG含量4、8周时分别为(5.13 ±0.32)mg/g、(5.37±0.12)mg/g.结论 以BMSCs作为种子细胞可于体内构建特定形态、组织学良好的Medpor-软骨复合体.     

构建带内支撑组织工程睾丸假体的实验研究

目的探讨以医用多孔高密度聚乙烯(high-density polyethylene,HDPE,商品名为Medpor)为内支撑,外裹聚羟基乙酸(polyglycolic acid,PGA)的支架,接种软骨细胞后,于裸鼠体内构建组织工程睾丸假体的可行性及特点。方法取新生雌性长枫杂交仔猪1只,取四肢大关节表面软骨,制备密度为5×107/ml的软骨细胞悬液。将其接种于长短半径分别为6mm和4mm的Medpor,外裹2mm PGA的支架,体外培养2周。取4周龄BALB/C裸鼠10只,随机分为两组(n=5)。实验组:采用制备的细胞-支架复合物植入裸鼠背部皮下;对照组:采用Medpor-PGA复合支架植入裸鼠皮下。8周后处死裸鼠取材,行大体观察、组织学及免疫组织化学观察。结果大体观察:实验组标本形状与体积未发生变化,色泽及弹性与正常软骨相似,软骨与Medpor接合紧密;对照组无软骨形成,外包裹纤维样组织。实验组:HE染色见软骨内存在大量成熟的软骨陷窝,无血管长入,部分PGA尚未降解完全;甲苯胺蓝染色示细胞外基质具有异染特性;藏红花染色示基质中大量蛋白聚糖沉积;型胶原免疫组织化学染色呈强阳性表达。对照组:Medpor被大量富含血管的纤维组织包裹。结论利用Medpor与PGA复合的支架,接种软骨细胞后,可于体内构建出具有预构睾丸形态且组织学良好的Medpor-软骨复合体。     

兔再造阴茎模型内构建组织工程化阴茎假体的实验研究

目的 探讨于家兔再造阴茎模型内构建带内支撑组织工程化阴茎假体的可行性及其特点.方法 取成年兔耳软骨分离培养软骨细胞,制备以医用多孔高密度聚乙烯(HDPE)为内支撑,外裹聚羟基乙酸(PGA)的支架(直径3mm、长20mm柱状HDPE,外裹厚2 mm的PGA),细胞-支架复合物体外培养4周.以兔一侧腹壁浅血管为轴型血管的皮瓣形成再造阴茎模型,将经体外培养的细胞-支架复合物分别植入兔即时再造的阴茎模型内和对侧腹壁皮下,在对侧腹壁下同时植入一单纯支架.将植入体分为A组:细胞-支架复合物植入再造阴茎模型内;B组:细胞-支架复合物植入腹壁下;C组:单纯支架植入腹壁下.术后3个月将兔处死取材,标本行大体观察、组织学、组织化学、免疫组化、糖胺聚糖(GAG)含量及生物力学检测.结果 A组与B组取材标本的体积、外形与植入时相近,外层新生组织与内部HDPE结合紧密,经HE染色、Safranin O染色、Masson trichrome染色和Ⅱ型胶原免疫组化检测证实,新生组织为软骨组织,且分布相对均一完整,纤维组织长人较少,少量炎性细胞浸润;C组无软骨组织形成.A组与B组标本各项检测指标(湿重、GAG含量、抗压强度、弹性模量)差异无统计学意义(P>0.05).结论 自体软骨细胞接种于HDPE-PGA复合支架,经体外培养4周后,植于家兔即时再造阴茎模型内继续培养,可成功地构建出具有预制形态、体积、结构与组织学良好的软骨-HDPE复合体(阴茎假体).     

膨体聚四氟乙烯（ePTFE）内支撑组织工程软骨的构建实验研究

目的：探讨骨髓间充质干细胞和软骨细胞混合培养体外构建ePTFE软骨复合体的可能性。方法：实验组：分离、获取、扩增兔骨髓间充质干细胞和软骨细胞,二者按7：3比例混和,接种到以膨体聚四氟乙烯（ePTFE）为内支撑外裹聚羟基乙酸（PGA）支架上,体外培养8周后,行大体、组织学、Ⅱ型胶原免疫组织化学和生物化学检测。对照组：利用实验组支架单纯接种骨髓间充质干细胞行体外培养。结果：实验组：体外培养8周后形成形态良好的软骨样组织复合体,组织学可见成熟软骨陷窝、异染基质、Ⅱ型胶原表达阳性。对照组：无软骨形成。结论：兔骨髓间充质干细胞和软骨细胞混合培养,可以在体外构建出特定形状、结构组织学良好的ePTFE软骨复合体。     

以β-磷酸三钙多孔陶瓷为载体建造组织工程化人工软骨

目的 通过将软骨细胞接种到三维多孔β－磷酸三钙（β－TCP）陶瓷支架材料上,探讨以β－TCP为载体建造组织工程化软骨的可行性。方法 将β－TPC多孔陶瓷加工成圆片状,并将其作为构建人工软骨的细胞支架。在支架材料上接种从兔关节软骨分离的软骨细胞,将细胞－陶瓷复合体置于旋转细胞培养系统（RCCS）内,培养1周后,将其移植到裸鼠背部皮下。植入术后4,8,16周取材,进行大体观察、组织学及组织化学等观察。结果 复合体体内移植后,在裸鼠皮下有新生软骨形成,形成的软骨基本保持了支架材料原有形态。结论 β－TPC多孔陶瓷是软骨细胞较适宜的贴附基质,以其为支架能够在体内建造出具有精确解剖形状的人工软骨。     

软骨细胞与脂肪基质细胞共培养体外构建软骨的初步研究

目的 探讨软骨细胞与脂肪基质细胞(adipose-derived stromal cells,ADSCs)共培养体外构建软骨的可行性,并阐明软骨细胞提供的软骨微环境能否诱导ADSCs向软骨细胞分化并形成软骨组织.方法 分别培养扩增人ADSCs与猪耳软骨细胞,将2种细胞按7:3(ADSCs:软骨细胞)比例混匀,以5.0×107/ml的细胞终浓度接种于聚羟基乙酸/聚乳酸(PGA/PLA,直径8 mm,高2 mm)支架作为共培养组,以相同终浓度的单纯软骨细胞和单纯ADSCs分别接种相同支架作为阳性对照组及阴性对照组,以30％上述浓度(1.5×107/ml)的单纯软骨细胞接种作为低浓度软骨细胞对照组.每组各接种6例标本,每例接种细胞悬液200μl.全部标本均于体外培养8周时取材,通过大体观察、组织学、免疫组化及湿重、蛋白多糖定量检测等方法对新生软骨进行初步评价.多样本t检验统计分析各组湿重及蛋白多糖含量差异.结果 各组细胞均与材料粘附良好.共培养组及阳性对照组体外培养8周时基本保持了复合物初始大小和形状,大体观察2组均形成了较成熟软骨组织,组织学显示大量软骨基质和软骨陷窝形成,免疫组化显示软骨特异性细胞外基质Ⅱ型胶原分泌.定量测定结果表明,共培养组的平均湿重为(174±12) mg,平均蛋白多糖含量为(7.6±0.4) mg,两者分别达到阳性对照组的75％ (P＜ 0.01)和79％ (P＜ 0.01).阴性对照组(单纯ADSCs组)明显皱缩变形,组织学未见成熟软骨陷窝.低浓度软骨细胞组明显变薄,新生软骨平均湿重为(85 ±5) mg,是阳性对照组的37％ (P＜ 0.01),只在局部形成了不连续的软骨组织.结论 软骨细胞与ADSCs共培养能够在体外构建较成熟的软骨组织,软骨细胞能够诱导ADSCs成软骨分化及体外形成软骨组织.     

同种异体软骨细胞与自体骨髓基质干细胞混合共培养构建软骨皮下移植的可行性研究

背景：软骨组织工程的种子细胞问题是目前研究的热点和难点,如何找到一种既能够避免对自体软骨进行取材又能够达到稳定软骨构建目的的方法呢？本研究尝试利用少量同种异体羊软骨细胞作为软骨诱导微环境提供者,与扩增后的羊自体BMSC混合共培养并植入皮下环境,探讨利用同种异体软骨细胞共培养构建软骨皮下移植的可行性。方法：本实验对山羊软骨细胞和BMSC分别进行取材和分离培养扩增,并将以上细胞分为以下四组进行混合并接种在PGA支架材料上：A组：100%自体软骨细胞;B组：30%自体软骨细胞＋70%自体BMSCs;C组：30%同种异体软骨细胞＋70%自体BMSCs;D组：100%同种异体软骨细胞。经过体外构建6周后植入羊皮下进行体内构建12周,对所形成的组织块进行大体观察和组织学染色等评价。结果：自体软骨细胞组和自体软骨细胞混合自体BMSC组皮下移植后可见成熟软骨组织形成,但同种异体软骨细胞参与的两组（包括同种异体软骨细胞混合自体BMSC的实验组和单纯异体软骨细胞组）在皮下环境中都因为较强的免疫反应未能形成软骨组织。结论：同种异体软骨细胞以及PGA支架材料的存在对于组织工程软骨在羊皮下环境的构建有负面影响。     

组织工程骨软骨复合体修复犬膝关节负重区骨软骨缺损的实验研究

目的 观察以骨软骨支架复合骨髓基质干细胞(bone-fflarrow mesenchymal stem cells,BMSCs)修复犬膝关节负重区骨软骨缺损的疗效.方法 利用软骨细胞外基质作为软骨支架部分,以脱细胞骨作为骨支架部分,采用相分离技术制备骨软骨双相支架,将成软骨诱导的BMSCs种植到双相支架上构建组织工程骨软骨复合体,并以此复合体修复犬膝关节股骨髁负重区骨软骨缺损,分为细胞-双相支架组(实验组)和单纯支架组(对照组).分别在术后3和6个月时取材,根据大体、组织学、Micro-CT等检测结果进行半定量或定量评估.结果 大体及组织学评价表明:同一时间点实验组的修复效果优于对照组,且实验组在术后6个月时的修复效果优于其术后3个月时,两项差异均有统计学意义;而对照组小同时间点修复效果的差异无统计学意义.Micro-CT检测结果表明实验组与对照组软骨下骨均得到重建,两者的差异尤统计学意义.结论 骨软骨双相支架复合成软骨诱导的BMSCs能成功修复犬膝关节负重区的骨软骨缺损,其修复效果明显优于单纯支架植入组.     

The Metabolism of Vitamin D3 in Response to Testosterone

Summary : Sex steroids were suggested as regulators of vitamin D metabolism. While considerable data is available regarding interaction between estradiol and vitamin D, very little is known about interactions between testosterone and vitamin D. A similar gap exists with regard to the involvement of the vitamin D endocrine system in the pathogenesis of the female versus the male osteoporosis syndrome. In the present study we studied the effect of long-term treatment with testosterone on the metabolism of vitamin D in vitamin D₃ replete sexually immature male chicks. We were able to show that under this treatment, circulating levels of 1,25-dihydroxy vitamin D₃ (1,25(OH)₂D₃) are significantly reduced, but intestine and bone concentrations are significantly increased. The increased concentration of 1,25(OH)₂D₃ in bone was accompanied by an increase in the ash content of this tissue. The reduction in serum 1,25(OH)₂D₃ was not dependent on reduced activity of the renal 25-hydroxy vitamin D₃ - 1α - hydroxylase. Based on these findings it is proposed that testosterone is involved in the stimulation of the biological response to vitamin D in the classical target-organs, such as intestine and bone, and this observation may provide partial explanation to the pathogenesis of osteoporosis in hypogonadal men.     

Postexercise albuminuria in children with different duration of type-1 diabetes mellitus

. About 30% of diabetic patients develop progressive renal failure. We studied albumin, IgG, and transferrin excretion during exercise in diabetic children without signs of nephropathy to investigate proteinuria under these conditions: 39 patients with insulin-dependent diabetes mellitus and 21 healthy children undertook a bicycle exercise test. Albuminuria measured by nephelometry was calculated as the albumin excretion rate (AER) and albumin-to-creatinine ratio before and after exercise. The diabetic group was divided into three subgroups according to disease duration (DI<5 years, DII 5 – 10 years, DIII>10 years). No significant difference in metabolic control (hemoglobin A_1c) was detected between the diabetic groups (median hemoglobin A_1c: DI 7.2%, DII 7.6%, DIII 8.6%). There was no increase in AER in the healthy children after exercise. Before exercise the diabetic groups had an AER similar to controls. No significant increase in albuminuria after exercise was seen in group DI. Both groups with a disease duration of more than 5 years had a significant increase in albuminuria [median before/after: DII 7.8/16.7 (P< 0.05), DIII 0/57.9 (P< 0.05) μg/min per 1.73 m²). Of these patients, 43% also had a measurable urinary excretion of IgG and transferrin, indicating structural glomerular damage. There was no correlation of albuminuria and parameters of metabolic control or renal function. We conclude that in diabetic children an exercise test unveils albuminuria in certain patients, while their AER may be normal at rest. Received September 22, 1995; received in revised form and accepted January 24, 1996     

何首乌提取物二苯乙烯苷对糖尿病大鼠肾脏氧化应激的影响

目的：何首乌提取物二苯乙烯苷（TSG）对糖尿病（DM）大鼠肾脏氧化应激干预机制。方法：采用左侧肾结扎术加尾静脉注射链脲佐菌素（STZ,40 mg/ kg）造糖尿病肾病（DN）大鼠模型,1周后随机分为模型组和 TSG 治疗组,治疗组给予 TSG（20 mg＆#183;kg -1＆#183;bw -1）腹腔注射,模型组接受等容量生理盐水。在6周末分别用代谢笼收集24 h 尿,计尿液总量后离心,用散色比浊法测定 MAU 和尿肌酐（Ucr）;HE 染色、电镜观察肾脏损伤后病理形态学变化;比色法、黄嘌呤氧化酶法测MDA、Scr、SOD。结果：TSG 减轻了 DN 大鼠肾脏损伤后病理形态学变化、减少了 UAER、MAU/ Ucr、MDA 的含量,升高了 SOD的含量,差异均有统计学意义（P 〈0．05）。结论：TSG 能够减少 DN 大鼠尿微量白蛋白,对 DM 大鼠肾脏具有保护作用,其机制可能与 TSG 改善 DM 大鼠肾组织中氧化与抗氧化之间的失衡有关。     

The expanding repertoire of targets for immune checkpoint inhibition in bladder cancer: What lies beneath the tip of the iceberg,PD-L1

Over the last decade, a new understanding of tumor-immune system interplay has been ushered in, lead in large part by the discovery of immune checkpoints mediated through B7-CD28 family interactions. Therapeutic blockade of the PD-L1 immune checkpoint pathway has already shown great success as a cancer immunotherapy for advanced urothelial carcinoma, leading to durable clinical remissions in an otherwise incurable disease. There are newly described members of the B7-CD28 family including B7-H3, B7x, and HHLA2. These ligands are thought to play an essential role in suppressing T-cell response, leading to immune tolerance of tumors. This feature makes them attractive targets for novel immunotherapy treatment paradigms. Here, we review the literature of current strategies and future directions of immune checkpoint blockade therapy for bladder cancer.     

Recent developments in the management of bladder cancer: Introduction

An upsurge of advances in the management of bladder cancer has rapidly occurred over the past 2 years. In this issue, recent developments in the management of bladder cancer will be discussed, including the emerging role of immunotherapy, biomarkers, and advanced imaging.     

Identification of men with low-risk biopsy-confirmed prostate cancer as candidates for active surveillance

Background

A combined clinical cell-cycle risk (CCR) score that incorporates prognostic molecular and clinical information has been recently developed and validated to improve prostate cancer mortality (PCM) risk stratification over clinical features alone. As clinical features are currently used to select men for active surveillance (AS), we developed and validated a CCR score threshold to improve the identification of men with low-risk disease who are appropriate for AS.

Trends in management of the small renal mass in renal transplant recipient candidates: A multi-institutional survey analysis

Introduction

Patients with end-stage renal disease are under increased risk for renal cell carcinoma development, and radical nephrectomy is the preferred treatment in this setting. Owing to the increased surgical morbidity and mortality, active surveillance (AS) may be a valid option for treatment of small renal masses (SRM). As there is a lack of high-level evidence for treatment recommendations, we performed a survey analysis to analyze the treatment patterns of transplant surgeons.