供体凋亡脾细胞输注诱导大鼠胰岛移植免疫耐受的研究

目的 探讨供体凋亡细胞输注诱导大鼠胰岛移植免疫耐受的可行性.方法 应用链脲佐菌素(STZ)制备SD大鼠糖尿病模型,以直线加速器照射诱导供体Wistar大鼠脾细胞凋亡.将糖尿病SD大鼠分为4组,分别经阴茎背静脉输注Hanks液、供体正常脾细胞、供体凋亡脾细胞、供体坏死脾细胞,7天后于肾包囊进行同种异体胰岛移植,测定血糖变化,观测胰岛移植物存活时间,并通过混合淋巴细胞反应(MLR)观察移植耐受的状态.结果 预输注供体凋亡脾细胞能显著延长同种异体胰岛移植物存活时间(中位存活时间达31天),并使受体鼠对供体鼠的MLR明显减弱.结论 供体凋亡细胞输注能够诱导同种异体大鼠胰岛移植免疫耐受.     

负载猪源性CTLA4Ig的猪胰岛细胞阻断直接识别途径在异种移植中的作用

目的：探讨负载供体源性CTLA4Ig的胰岛细胞通过阻断直接识别途径,诱导异种胰岛细胞抑制免疫耐受的可行性及有效性。方法：使用本实验小组前期构建的携带供体源性（猪）CTLA4-Ig的腺病毒载体（Adv-pCT-LA4-Ig）转染猪胰岛细胞,植入糖尿病大鼠肾包膜下,建立猪-大鼠异种胰岛移植模型,监测移植术后大鼠血糖变化及IL-4、γ-IFN的水平变化,检测移植物中pCTLA4-Ig、猪源性胰岛素（p-Insulin）表达情况。结果：1.大鼠血糖在移植术后即开始明显下降,于第3 d左右降至正常,三组大鼠血糖分别在移植术后6、7、35 d左右开始升高;2.三组大鼠术后胰岛平均存活时间为7.43±1.72 d、7.22±1.72 d、34.50±4.14 d,实验组胰岛细胞存活时间较对照组明显延长（P〈0.01）;3.细胞因子变化：（1）γ-IFN：对照组在移植术后第1 d开始轻度升高,于第3 d后开始明显升高,第7 d达到高峰,此后缓慢升高;实验组在移植术后第1～28 d波动较小,同术前相比无明显变化,在移植后第28 d开始上升,第35 d开始急剧上升;（2）IL-4：实验组在移植术后第1 d开始下降,第7 d达到谷值;对照组在移植术后第3～5 d轻微上升,在移植后第28 d开始下降至移植术前水平,第35 d开始急剧下降;4.病理学检查：实验组移植物无明显破坏,炎细胞浸润不明显;免疫组化可见大量pCTLA4-Ig、p-Insulin表达。结论：转染供体源性CTLA4-Ig的胰岛细胞通过阻断直接识别途径,可诱导异种胰岛细胞产生较长时间的免疫耐受。     

hGLP-1类似物基因转染对糖尿病大鼠血糖、血清胰岛素水平及胰岛细胞的影响

目的 研究尾静脉注射人胰高血糖素样肽-1(hGLP-1)类似物基因(2×Val~2-hGLP-1)重组表达质粒对糖尿病模型大鼠血糖、血清胰岛素水平及胰岛细胞的影响.方法 建立链脲佐黹素(STZ)诱导SD糖尿病大鼠模型,随机分为含有hGLP-1类似物基因的重组质粒(plRFLS2-FGFP/Val~2-hGLP-1)转染组、空质粒(pIRES2-EGFP)转染组、糖尿病模型对照组,每组8只.另取8只未经处理的SD大鼠作为正常对照组.重组质粒转染组和空质粒转染组分别通过尾静脉注射含相应质粒(110μg/只)的溶液,糖尿病模型对照组和正常对照组给予等量的生理盐水.实验动物共干预30d,实验结束后,观察各组大鼠血糖及血清胰岛素水半的变化,采用糖耐量及胰岛素耐量实验检测大鼠胰岛素敏感性然后处死动物,HE染色观察胰岛细胞病变情况,免疫组化法分析胰岛素分泌情况.结果 与正常对照组、糖尿病模型对照组和空质粒转染组比较,重组质粒转染组血糖水平明显降低(P<0.01),血清胰岛素水平显著升高(P<0.01).基因治疗改善了糖尿病大鼠的糖耐量,降低了胰岛素抵抗,提高了机体对胰岛素的敏感性(P<0.01).空质粒转染组与糖尿病模型对照组比较,以上指标的差异均无统计学意义(P>0.05).同时,重组质粒转染组的胰岛素分泌水平提高,与糖尿病模型对照组比较,胰岛细胞病变程度明显改善.结论 hGLP-1类似物基因转染可促进胰岛细胞增殖,提高胰岛素敏感性,从而显著降低糖尿病大鼠的血糖水平,并改善胰岛组织病变程度.     

间充质干细胞联合骨髓细胞输注诱导胰岛移植免疫耐受的研究

目的观察间充质干细胞(MSC)联合骨髓细胞(BMC)输注对同种异体小鼠胰岛移植嵌合状态及胰岛移植物存活时间的影响。方法C57BL/6小鼠和BALB/C小鼠分别作为供、受体。应用链脲佐菌素制备BALB/C小鼠糖尿病模型,将分离纯化的C57BL/6小鼠胰岛移植到上述糖尿病模型小鼠的肾包囊下,用抗CD154单抗进行受体预处理。将接受胰岛移植的25只BALB/C受体鼠随机分为A组(单纯胰岛移植);B组(供体MSC悬液0.5ml输注);C组(供体BMC悬液0.5ml输注);D组(供体BMC和MSC各0.5ml输注);E组(供体BMC和第三品系的KM小鼠MSC各0.5ml输注),每组5只,所有细胞在胰岛移植后经尾静脉输注。比较以上各组供体细胞嵌合率(DC)和胰岛移植物存活时间的差异。结果在胰岛移植后第30天,MSC联合BMC输注的D、E两组与单纯BMC输注的C组比较,其DC显著升高(P<0.01);胰岛存活时间亦显著延长[(77.0±7.7d)和(61.0±2.2d)vs(53.0±16.4d)(P<0.01)]。胰岛移植后第60天,D组与E组比较,DC维持在更高水平,且胰岛移植物存活时间更长(P<0.05)。结论MSC联合BMC输注比单纯BMC输注能够维持更长时间的混合嵌合状态,并延长胰岛移植物存活时间;供体来源的MSC输注比非供体来源的MSC输注更有效。     

模拟微重力条件下大鼠胰岛培养及体内移植的实验研究

目的研究经模拟微重力条件培养的胰岛是否能够降低胰岛移植过程中引发的免疫排斥反应，进而延长胰岛的体内存活时间。方法分离新鲜胰岛，应用培养皿对胰岛进行静态培养，同时应用旋转式生物反应器对胰岛进行三维立体培养。应用吖啶橙一碘化丙啶（AO-PI）染色和葡萄糖刺激实验对3种条件下培养得到的胰岛进行生物学活性研究。2000当量胰岛在旋转式生物反应器中培养5d后，将其移植入经链脲佐菌素（STZ）处理的糖尿病模型SD大鼠。肾被膜下作为实验组，以新鲜分离的胰岛和静态培养5d的胰岛移植入经STZ处理的糖尿病模型SD大鼠肾被膜下作为对照组。在不同时间点上，检测各组SD大鼠血糖变化情况；对各移植组中大鼠进行糖耐受实验。切除。肾被膜下的移植物组织作为标本，进行苏木精一伊红（HE）染色及胰岛素组织化学染色检测。结果体外检测结果表明，与新鲜分离的胰岛和静态培养的胰岛相比，微重力条件下培养的胰岛仍保持良好的胰岛素合成及分泌功能。体内实验表明，微重力条件下培养的胰岛体内移植组的血糖保持正常的时间明显高于接受静态培养的胰岛和新鲜分离的胰岛移植组。将微重力培养条件下胰岛移植大鼠后，取。肾被膜下移植区组织标本进行HE染色，未见明显淋巴细胞浸润，胰岛素免疫组织化学均可见阳性细胞。而新分离的胰岛以及静态培养的胰岛移植后，大鼠。肾被膜下移植物组织标本可见淋巴细胞浸润，移植物的厚度明显变小。结论胰岛在微重力条件下培养可降低移植引起的免疫排斥反应，进而延长移植物体内存活时间。     

口服抗原对同种异体移植免疫反应的影响

将Wistar大鼠预先喂养同种异基因SD大鼠脾细胞7天，然后接受SD大鼠的皮肤移植，7天后，再接近SD大鼠的心脏移植，观察口服抗原后体外混合淋巴细胞培养（MLC）、体内迟发型超敏反应（DTH）以及心脏存活时间。口服抗原后，体外MLC及体内DTH反应均出现明显的抗原特异性降低，口服抗原组大鼠并接近供者皮肤致敏后的心脏移植存活时间达到7天，与对照组未致敏鼠的心脏移植存活时间一致；而未口服抗原组接受皮肤致敏后的心脏移植存活时间不超过2天。提示口服抗原可以使异基因抗原的特异性免疫应答降低，延长移植存活时间，阻皮肤移植物的事先致敏反应的发生，并使加速排斥反应时间明显延迟。     

糖尿病犬经肝动脉肝内移植微囊化猪胰岛细胞的安全性和有效性

目的评估移植于I型糖尿病犬肝脏中的微囊化新生猪胰岛细胞(NPI)的生物相容性、免疫学特性及生理学特性。方法I型糖尿病犬分为A、B两组,每组15只,A组每只犬分别经肝动脉灌注微囊化新生猪胰岛细胞40～60万个,B组每只犬分别经肝动脉灌注未微囊化新生猪胰岛细胞40～60万个,两组动物移植后均不使用免疫抑制治疗。移植前后分别测量移植受体的肝脏功能及淋巴细胞CD4 /CD8 比值。移植6个月后所有移植受体的肝脏均进行病理学检查。结果移植后A组外源性胰岛素用量从移植前的22 u逐渐降至5 u,B组所需外源性胰岛素从移植前的24 u下降至10 u。移植后2～3周B组胰岛素用量恢复到移植前的水平,而A组的部分动物的胰岛素剂量继续减至8 u。B组受体移植后血的CD4 较移植前升高,而A组的CD4 和CD8 细胞移植后无明显变化。移植后所有受体的肝功能及组织结构未见异常。结论微囊化的新生猪胰岛细胞在受体犬的肝脏中有很好的生物相容性。微囊可以延长移植物的存活,且异种移植微囊化的新生猪胰岛细胞能够纠正糖尿病犬的高血糖状态。     

人脑源性神经营养因子基因转染神经干细胞治疗大鼠视神经损伤

目的 探讨转染人脑源性神经营养因子(brain-derived neurotrophic factor,hBDNF)-绿色荧光蛋白(GFP)基因神经干细胞体内移植后在视神经损伤大鼠视网膜内的存活、迁移和分化情况,以及其对视神经损伤后视网膜神经节细胞(retinal ganglion cells,RGCs)存活的影响.方法 (1)取30只SD大鼠制作右眼视神经部分损伤模型,采用随机数字表法随机均分成两组,分别于玻璃体腔内移植GFP基因转染神经干细胞(GFP-NSCs)、hBDNF和GFP双基因转染神经干细胞(hBDNF-GFP-NSCs).8周后处死大鼠,取右眼眼球做冰冻切片,然后分别用胸腺细胞表面糖蛋白(Thy1.1)、胶质纤维酸性蛋白(GFAP)、β-微管蛋白(β-tubulin)、视紫红质(rhodopsin)抗体进行免疫标记,观察移植细胞的迁移分化情况.(2)取30只SD大鼠制作右眼视神经部分损伤模型,随机均分为三组:损伤组、GFP组、BDNF组.损伤组大鼠玻璃体腔内注射等渗盐水,而GFP组、BDNF组则分别于玻璃体腔内移植GFP-NSCs和hBDNF-GFP-NSCs,每只眼球玻璃体腔内移植5×104个细胞.8周后处死大鼠,取右侧眼球视网膜铺片,行Thy1.1免疫荧光染色后,荧光显微镜下观察计数RGCs细胞数量.结果 (1)移植后,两组细胞均可在视网膜内存活、迁移,并产生细胞分化,两组细胞均可以分化成胶质细胞、神经元.移植hBDNF-GFP-NSCs组少量细胞分化为节细胞样细胞(Thy1.1+),而GFP-NSCs组未发现有分化为Thy1.1阳性细胞.(2)视网膜铺片Thy1.1免疫荧光染色后计数发现,移植hBDNF-GFP-NSCs的视网膜神经节细胞为(1461±154)个/mm2,明显多于移植GFP-NSCs组(1 244±187)个/mm2(P<0.05).结论 移植后的hBDNF-GFP-NSCs可以分化为神经元和神经胶质细胞,少数可分化为视网膜神经节样细胞.hBDNF-GFP-NSCs移植可以增加RGCs的存活数量,有助于视神经损伤后的修复.     

海藻酸钠-多聚赖氨酸-海藻酸钠微囊化胰岛细胞移植治疗糖尿病的实验研究

采用Ｓｕｎ海藻酸钠－多聚赖氨酸－海藻酸钠（ＡＰＡ）微囊制作技术,分别包裹大鼠胰岛和胰岛素分泌细胞系,移植于糖尿病小鼠腹腔。结果表明ＡＰＡ微囊化大鼠胰岛或胰岛素分泌细胞移植,均可使糖尿病小鼠血糖降低至接近正常水平达３周至１１０天;移植微囊无明显的组织学反应。证明该ＡＰＡ微囊化胰岛细胞移植具有较好的治疗效果,微囊具有较好的生物相容性和免疫隔离作用。为进一步发展生物型人工胰岛奠定了基础。     

胰岛移植的分子影像

对不稳定1型糖尿病而言,胰岛移植是使患者脱离胰岛素治疗的一种有效方法。传统影像检测不能有效地监测与评估体内移植的胰岛,分子影像学能实时对移植的胰岛移植物的存活状态及功能进行评估,从而为临床提供更有参考价值的、实时反映移植物状态的数据,对移植物的活力及功能进行活体评估,有利于胰岛移植的后续干预治疗,将有望推动胰岛移植临     

Stress MR imaging for evaluation of popliteal artery entrapment

The popliteal artery entrapment (PAE) syndrome has been recognized as a cause of arterial occlusion in young people. It is the result of an anomaly of the relationship between the popliteal artery and the gastrocnemius muscle. Eight young healthy volunteers (16 legs) and six patients (10 legs) with suspected PAE underwent magnetic resonance (MR) imaging. Gradient-echo images were obtained in axial planes with the leg at rest and during active plantar flexion against resistance. Imaging at rest allowed identification of PAE signs in only one leg, which had an anomalous medial course of the popliteal artery. In the other cases, only the stress technique was able to show signal loss in the popliteal artery due to muscular compression (two legs) or the presence of accessory muscle slip around the vessel (two legs), as confirmed at surgery. MR imaging is therefore a useful technique for the diagnosis of PAE because of its capability of combining information obtainable with other modalities.     

MR imaging of muscle and tender points in fibromyalgia

Fibromyalgia is a syndrome manifested by chronic, diffuse muscu-loskeletal aching and soreness, palpable muscle tender points, and other symptoms. Standardized clinical diagnostic criteria have recently been developed. Skeletal muscle has been postulated as the end organ in this disease. Biochemical, histologic, electromyographic, and conventional radiographic studies have demonstrated no definitive abnormality. This study sought to establish whether magnetic resonance (MR) imaging could demonstrate any abnormality in these patients. Eighteen patients were entered in the study, 14 of whom were able to complete their examinations. T1 -weighted, T2-weighted, gradient-echo, and STIR (short-tau inversion-recovery) sequences were performed in all patients, with selected patients examined with T1weighted, gadopentetate dimeglu-mine-enhanced sequences. The trapezius and suboccipital regions were imaged in patients who, clinically, had active fibro-myalgia. No abnormalities could be detected. The authors conclude that the conventional MR imaging used in this study was unable to depict any primary skeletal muscle abnormality in fibromyalgia.     

Uterine contractions: Possible diagnostic pitfall at MR imaging

A total of 206 nongravid patients with various gynecologic problems underwent pelvic magnetic resonance (MR) examinations that included both sagittal T2-weighted and contrast agent–enhanced T1-weighted images. MR images were retrospectively reviewed to identify changes in endometrial configuration on serial images obtained during the same MR examination. In 20 MR examinations (all in women of reproductive age), endometrial distortion due to myometrial bulging was noted on T2-weighted or contrast-enhanced T1-weighted images. It was absent on other MR images obtained at different times. Myometrial bulging exhibited low signal intensity in 18 examinations. The finding resembled adenomyosis or leiomyoma on T2-weighted or contrast-enhanced T1-weighted images. These results evidence the presence of transient myometrial bulging and transient low-intensity myometrium in the nongravid uterus. This phenomenon is thought to represent uterine contraction. Clinicians should be aware of the potential presence of transient low-signal-intensity myometrial bulging that could present diagnostic problems in the normal uterus.     

Controversies in cervical spine imaging in trauma patients

No area of emergency radiology has generated as much discussion in recent years as the subject of cervical spine imaging for trauma patients. This review will be in three parts. The first will examine the indications for cervical imaging and will focus on those factors that make patients at high risk or low risk for cervical injury. The second part will discuss the merits of radiography and computed tomography as the main screening diagnostic examination. In addition to the roles of each modality in the evaluation process, such factors as efficacy of diagnosis, time (duration) of study, and cost will be discussed. Finally, the third part will explore the methods currently employed to clear the cervical spine in comatose patients.Presented at the Annual Meeting of the American Society of Emergency Radiology, Las Vegas, Nevada, 22–25 October, 2003     

Brodie Abscess: MR imaging appearance in 10 patients

The magnetic resonance (MR) imaging features of Brodie abscess have not yet been fully evaluated. Ten patients with Brodie abscess, eight of long bone and two of vertebra, were studied with MR imaging. Long bone abscess had a characteristic “target” appearance with four layers: (a) a center with low signal intensity on T1-weighted images and high signal intensity on T2-weighted and STIR (short-inversion-time inversion recovery) images, (b) an inner ring isointense to muscle on T1-weighted images and with high signal intensity on T2-weighted and STIR images, (c) an outer ring hypoin-tense on all images, and (d) a peripheral halo hypointense on T1-weighted images. In six of eight cases, a soft-tissue mass was found. The two vertebral abscesses had a less specific appearance, with low signal intensity on T1-weighted images and high signal intensity on T2-weighted and STIR images. Only the peripheral halo was clearly identified in both cases.     

Dynamic contrast-enhanced MR imaging assessment of vascularized free fibular grafts

Magnetic resonance (MR) imaging may be a noninvasive method for assessing perfusion of vascularized bone grafts placed for treatment of avascular necrosis. One proximal femur of seven beagles was devascularized, with insertion of a vascularized fibular graft. MR imaging at 1 week (seven dogs) and 6 weeks (five dogs) after surgery included pre- and postcontrast spin-echo sequences, unenhanced twodimensional time-of-flight (TOF) vascular imaging, and dynamic gradient-echo imaging during infusion of gadolinium. Relative signal intensity values of selected regions obtained from the dynamic gradientecho images were plotted as percent enhancement versus time. In the operated hip, MR imaging did not show enhancement in six of seven femoral heads and greater trochanters at 1 week after surgery, with similar results after 6 weeks. MR imaging of fibular grafts 6 weeks after surgery showed an initial rapid increase in enhancement and a subsequent slower increase in five of five dogs, although no enhancement was seen in six of seven dogs at 1 week. These findings contrasted with a rapid initial increase in enhancement followed by slow decline in non-operated hips. Two-dimensional TOP imaging did not show the vascular pedicle of the graft in any dog. Findings of radionuclide bone scanning performed 1 week after surgery were consistent with devascularization of the operated femur and fibular graft. However, tetracycline distribution and histologic findings confirmed the viability of five of five grafts within the devascularized femurs 6 weeks after surgery. Thus, dynamic contrast-enhanced MR imaging at 6 weeks after surgery is valuable for assessing vascular bone graft perfusion, while similar imaging at 1 week may suggest otherwise.     

Right-side aortic arch with aneurysm of aberrant left subclavian artery: MR imaging appearance.

Reports of aneurysms of the subclavian artery in both normal and anomalous aortic arches have been rare. The authors describe a patient with a right-side aortic arch and an aneurysm of the aberrant left subclavian artery, which, to the authors' knowledge, is a previously unreported association. At presentation, the aneurysm appeared as a calcified left superior mediastinal mass. Magnetic resonance imaging enabled preoperative diagnosis and guided surgical planning.     

Adrenal tissue characterization with 0.5-T MR imaging: Value of T2*-weighted images

The authors investigated the value of magnetic resonance (MR) imaging at 0.5 T for distinguishing adrenal adenomas from adrenal metastases. The series included 23 adrenal adenomas (18 nonhyperfunctioning, five hyperfunctioning) and 23 adrenal metastases from various organs. Adrenal tumor–liver signal intensity ratios on T1-, T2-, and T2*-weighted images were calculated for adrenal tissue characterization. Adrenal adenomas were more precisely distinguished from adrenal metastases on T2*-weighted images (21 of 23, 91%) than on T2-weighted images (15 of 23, 65%). T1-weighted images were not useful for this distinction. In conclusion, T2*-weighted images were better than routine T2-weighted images for distinguishing adrenal adenomas from adrenal metastases. It can be postulated that the total signal intensity of adrenal adenomas, which contain some fat components, decreased on T2*-weighted images because of an out-of-phase effect.     

MR angiography of congenital heart disease: Value of segmented two-dimensional inflow technique and maximum-intensity-projection display

Magnetic resonance (MR) angiography of the cardiovascular system was evaluated in 41 patients with congenital heart disease by using a two-dimensional (2D) inflow technique based on a magnetization-prepared gradient-echo pulse sequence with segmented k-space data acquisition and electrocardiographic gating at 0.5 T. Inversion and saturation prepulses were used to suppress stationary tissue and enhance intravascular signal. Presaturation slabs were applied where certain vascular structures had to be suppressed. Sequence parameters were optimized by evaluating signal intensity and contrast characteristics for various flip angles and inversion and saturation delay times. The heart and intrathoracic vasculature were encompassed with 40–50 overlapping sections. Both 2D angiograms and maximum-intensity-projection images were evaluated. Combining data sets acquired in the sagittal and transverse orientations provided the most satisfactory information about the pulmonary arteries. The highest signal-to-noise ratios were obtained with a flip angle of 65° and short prepulse delay times. Two-dimensional MR angiography can provide useful diagnostic information but requires a thorough understanding of in-plane and hemodynamically induced signal intensity changes.