miR-27a promotes cell proliferation and metastasis in renal cell carcinoma

miR-27a has been reported to exhibit abnormal expression in renal cell carcinoma (RCC), but the role of miR-27a in RCC remains unknown. In our study, up-regulation of miR-27a was validated by Real-time PCR analysis in 133 RCC samples. Overexpression of miR-27a promoted cell migration, invasion and proliferation in vitro, while its low expression exerted opposite effects. Kaplan-Meier analysis demonstrated that the patients with high expression of miR-27a had a worse overall and relapse-free survivals compared with those with low expression of miR-27a. Cox proportional hazards analyses showed that miR-27a expression was an independent prognostic factor for RCC patients. Collectively, our findings illustrate the promoting-cancer effect of miR-27a in RCC, suggesting that miR-27a could be a potential therapeutic target for RCC. Additionally, Kaplan-Meier analyses and Cox proportional regression analysis suggest that miR-27a may be a potential biomarker for predicting the survival of RCC patients.     

阻断VEGFR-3表达对肿瘤细胞诱导的淋巴管内皮细胞增殖的影响

目的观察阻断VEGFR-3(F lt 4)表达对肿瘤细胞(前列腺癌细胞株PC3)诱导的淋巴管内皮细胞增殖的影响。方法实验分4组,第1组为对照组。每组有6孔,每孔具有相同细胞数2×105/m l,实验组每孔各加入试剂100 m l/L。第1组(对照组)加入淋巴管内皮细胞完全条件培养液(LEC)1 m l,第2组加入LEC 1 m l+兔血清100μl,第3组加入LEC 1 m l+PC3细胞上清100μl。第4组加入LEC+抗F lt 4抗体100μl。并于24、48、72、96 h观察各组淋巴管内皮细胞生长情况。各时间段计数第1~3组细胞数,第4组于72 h计数后,去除抗体,重新加入LEC+PC3细胞上清继续培养至120 h;除96 h外,其余各时间段均计数细胞数。比较各组细胞增殖情况。结果第1、2组淋巴管内皮细胞在加试剂后各时间段两组细胞数及形态无明显差别,第3组加入PC3细胞上清后,细胞数明显多于第1、2组。第4组加试剂后24、48、72 h细胞计数均少于前24 h。清除抗体后加入PC3细胞上清48 h计数细胞,细胞数仅略见增加。结论VEGF-C高表达的PC3细胞上清能显著刺激淋巴管内皮细胞增殖,阻断F lt4表达,可在一定程度上阻断PC3细胞上清促淋巴管内皮细胞增殖作用。     

Caveolin-1 promotes tumor cell proliferation and vasculogenic mimicry formation in human glioma

Vasculogenic mimicry (VM) plays an important role in human glioma progression and resistance to antiangiogenic therapy as a compensatory neovascularization mechanism in malignant tumors. Caveolin-1 (Cav-1) has been found to contribute to VM formation. However, it remains largely unknown whether Cav-1 expression correlates with VM in glioma. In this study, we examined CAV-1 expression levels and VM in human glioma cell lines and in 94 human gliomas with different grades of malignancy, and present Cox proportional hazards regression. The molecular role of Cav-1 in glioma cells was investigated using quantitative polymerase chain reaction (qRT-PCR) assays, western blotting, CCK-8 assays, and tubule formation assays. Cav-1 expression and VM formation were positively correlated with each other and both were closely associated with glioma development and progression as evidenced by the presence of cystic tumor, shortened survival time, and advanced-stage glioma in glioma patients with Cav-1 overexpression/increased VM formation. Cav-1 promoted U251 glioma cell proliferation and VM formation in a Matrigel-based 3D culture model. VM-associated factors including hypoxia-inducible factor 1α (HIF-1α) and p-Akt was significantly elevated by Cav-1 overexpression but suppressed by siCav-1 in U251 cells. Collectively, our study identified Cav-1 as an important regulator of glioma cell proliferation and VM formation, contributing to glioma development and progression.     

Increased HIF-1alpha expression in tumor cells and lymphocytes of tumor microenvironments predicts unfavorable survival in esophageal squamous cell carcinoma patients

The expression of hypoxia-induced factor (HIF)-1α is up-regulated in tumor microenvironments under hypoxia condition. However, the prognostic significance of HIF-1α in esophageal squamous cell carcinoma (ESCC) is still elusive. We measured the HIF-1α expression by immunochemistry in tumor specimens from 136 resected ESCC; in the current study, the HIF-1α expression in tumor cells was significantly associated with tumor stage (P = 0.003) and lymph node metastasis (P = 0.006); whereas the HIF-1α expression in tumor-infiltrating lymphocytes (TILs) had no relationship with patients’ clinicopathological parameters. Patients with high HIF-1α expression in tumor cells or in TILs showed worse survival related to those with low HIF-1α expression. Multivariate analysis demonstrated that expression of HIF-1α in TILs was an independent factor for DFS (P = 0.007) and OS (P = 0.013). Additionally, the expression of HIF-1α in tumor cells was an independent factor for DFS (P = 0.037) and OS (P = 0.033) in locoregional ESCC patients, whereas the expression of HIF-1α in TILs was an independent factor for DFS (P = 0.048) and OS (P = 0.039) in metastatic ESCC patients. Correlation analysis revealed that expressions of HIF-1α in tumor cells and in TILs were positively correlated, and patients with combined high HIF-1α in both tumor cells and TILs had the worst survivals (P < 0.05). These findings suggest that the HIF-1α expressions in different cell populations of ESCC microenvironments have different clinical relevance and prognostic impact on patients.     

Plasma level and tissue expression of vascular endothelial growth factor in renal cell carcinoma: a prospective study of 50 cases

Vascular endothelial growth factor (VEGF) is the major factor involved in angiogenesis. Although it is known that one of the functions of VEGF is to regulate neovascularization in renal cell carcinomas, the relationship between the production of VEGF in tumor tissue and its concentration in blood has not yet been studied. The aims of this study were to determine, in a series of conventional renal cell carcinoma (CRCC) cases, (1) VEGF expression and VEGF pattern in tumor cells, (2) the relationship between VEGF expression/pattern and VEGF levels in plasma (pVEGF), and (3) the association with usual clinical and pathologic prognostic factors. Fifty patients operated on for CRCC by radical nephrectomy were included. Clinical and histologic parameters were studied. VEGF expression and VEGF pattern in tumor cells was immunohistochemically recorded. pVEGF levels and platelet count were analyzed in relation to clinical and histologic parameters. Intratumoral VEGF expression associated with a cytoplasmic VEGF pattern was significantly higher in patients with high pVEGF levels (P = .01). Both VEGF expression and pVEGF levels were significantly correlated with Fuhrman grade (P = .002 and P = .01, respectively) and tumor stage (P = .006 and P = .008, respectively). In addition, VEGF expression was also correlated with tumor necrosis (P = .001) and progression (P = .001). We demonstrated that in CRCC with tumor necrosis, VEGF expression, pVEGF levels, and platelet count were significantly higher than in CRCC with no tumor necrosis (P = .001, P = .03, and P = .001, respectively). Our results revealed that cytoplasmic VEGF expression and pVEGF levels are associated with usual prognostic factors and progression in CRCC, which may allow VEGF to be used as a prognostic marker for CRCC, especially in patients with VEGF-targeted therapy.     

内皮祖细胞对小鼠肾癌血管新生化的影响

目的 检测内皮祖细胞（EPCs）在小鼠肾癌（RCC）中的分布特征,探讨EPCs在肾癌新生血管化中的作用。方法 雄性BALB／c裸鼠70只按数字表法随机分为A组（35只）、B组（35只）,另取6只作为对照组（C组）。A组小鼠右肾下极移植人肾癌细胞,B组为假手术,C组不做任何处理;于实验后21、28、35、42、49d,A、B、C组各时间点分别处死7只、7只和1只小鼠,收集标本评估C组右侧正常肾组织（NT）、B组右肾损伤肾组织（ST）、A组的癌组织（1Tr）和癌旁组织（AT）中EPCs水平及分布,以及微血管密度（MVD）、血管内皮生长因子（VEGF）、基质细胞衍生因子（SDF-1）、内皮生长因子受体2（FLK）、趋化因子受体（CXCR4）等mRNA表达。结果 流式细胞仪检测显示,AT中EPCs水平在28d后逐渐升高,明显高于TT、NT及ST中的表达（P〈0．05）;TT中EPCs水平则先升高以后逐渐降低,且明显高于NT中表达（P〈0．05）。免疫荧光检测EPCs主要聚集于AT;免疫组织化学检测AT中MVD明显高于TT和NT,而TT中MVD则低于NT（P〈0．05）;AT中VEGF、FLK、SDF-1和CXCR4 mRNA水平对比,TT和NT明显升高（P〈0．05）。结论 EPCs可通过分泌促血管内皮生长因子和参与血管形成来参与肿瘤新生血管化,并以此促进肾癌的进展。     

Expression of vascular endothelial growth factor by human renal cancer cells enhances angiogenesis of primary tumors and production of ascites but not metastasis to the lungs in nude mice

We determined the role that vascular endothelial growth factor (VEGF), also known as vascular permeability factor (VPF), plays in the progression of human renal cell cancer in nude mice. Low metastatic and low VEGF/VPF-expressing human renal cancer cells SN12C were transfected with the VEGF165 cDNA or plasmid alone as control. VEGF165-transfected SN12C cells produced large amounts of biologically active VEGF in culture that did not affect cell doubling time or confluence. Subsequent to implantation into the renal subcapsule of nude mice, the VEGF165-transfected SN12C cells produced fast-growing (PCNA labeling), large tumors that expressed high levels of VEGF/VPF and were well vascularized (CD3-positive vessels). The tumors produced hyperpermeability of peritoneal blood vessels (Evans blue dye-leak assay), bloody ascites, and short survival time. Parental or control transfected SN12C cells produced less vascularized, slower growing tumors with no ascites. Regardless of in vivo expression level of VEGF, the incidence of spontaneous lung metastasis was low, suggesting that in itself, the expression of VEGF/VPF by renal cancer cells is not sufficient to produce metastasis. This revised version was published online in July 2006 with corrections to the Cover Date.     

In vivo and in vitro characteristic of HIF-1α and relative genes in ischemic femoral head necrosis

Background: Legg-Calvé-Perthes Disease (Perthes’ disease) is a childhood hip disorder initiated by ischemic necrosis of the growing femoral head. So far, the etiology and pathogenesis of Perthes’ disease is poorly understood. Materials and methods: Avascular osteonecrosis rat model was established to mimic the pathophysiological changes of femoral head necrosis. The chondrocytes of newborn Sprague-Dawley rats were isolated and cultured in hypoxic and normoxic condition. The expression characteristic of the hypoxia-inducible factor-1 alpha (HIF-1α) was evaluated both in vivo and in vitro models. Vascular endothelial growth factor (VEGF) and apoptotic genes in chondrocytes treated with normoxia and hypoxia were also studied. Results: HIF-1α expression increased greatly after ischemic operation and kept at relative high level in the arthromeningitis stage and declined in the stages of osteonecrosis and reconstruction. The HIF-1α mRNA levels of chondrocytes incubated at hypoxia were significantly higher than the cells treated with normoxia at 24 and 72 hours. Hypoxia inhibited VEGF expression; chondrocytes could oppose this inhibition manifested by the increasing of VEGF mRNA level after 72 hours hypoxia. The expression of apoptotic genes, Casp3, Casp8 and Casp9, elevated in chondrocytes after hypoxia with time differences. Conclusion: Hypoxia might be an etiological factor for femoral head necrosis, HIF-1α, VEGF as well as apoptotic genes participated the pathophysiological process of ischemic osteonecrosis.     

Distinct patterns of ALDH1A1 expression predict metastasis and poor outcome of colorectal carcinoma

Purpose: Aldehyde dehydrogenase 1A1 (ALDH1A1) has been proposed as a candidate biomarker for colorectal carcinoma (CRC). However, the heterogeneity of its expression makes it difficult to predict the outcome of CRC. The aim of this study was to evaluate the diagnostic and prognostic value of this molecule in CRC. Methods and Results: In this study, we examined ALDH1A1 expression by immunohistochemistry including 406 cases of primary CRC with corresponding adjacent mucosa, with confirmation of real-time PCR and Western blotting. We found that the expression patterns of ALDH1A1 were heterogeneous in the CRC and corresponding adjacent tissues. We defined the ratio of ALDH1A1 level in adjacent mucosa to that in tumor tissues as R_A/C and found that the capabilities of tumor invasion and metastasis in the tumors with R_A/C < 1 were significantly higher than those with R_A/C ≥ 1. Follow-up data showed the worse prognoses in the CRC patients with R_A/C < 1. For understanding the underlying mechanism, the localization of β-catenin was detected in the CRC tissues with different patterns of ALDH1A1 expression from 221 patients and β-catenin was found preferentially expressed in cell nuclei of the tumors with R_A/C < 1 and ALDH1A1^high expression of HT29 cell line, indicating that nuclear translocation of β-catenin might contribute to the increased potentials of invasion and metastasis. Conclusion: Our results indicate that R_A/C is a novel biomarker to reflect the distinct expression patterns of ALDH1A1 for predicting metastasis and prognosis of CRC.     

MiRNA-15a inhibits proliferation,migration and invasion by targeting TNFAIP1 in human osteosarcoma cells

Recent data strongly suggest the important role of miRNAs in various cancer-related processes. Osteosarcoma is the most common type of primary malignant bone tumor and is characterized by complex genetic changes and resistance to conventional treatments. In this study, the role of miRNA-15a (miR-15a) in the progression and metastasis of osteosarcoma was investigated. The result demonstrated that the expression of miR-15a was down-regulated in osteosarcoma tissues and cell lines as compared with that in adjacent non-neoplastic bone tissues and the osteoblastic cell line. In functional assays, miR-15a inhibited cell proliferation, migration and invasion in U2OS and MG-63 cells. Meanwhile, bioinformatic analysis combined with experimental confirmation demonstrated that tumor necrosis factor; α-induced protein 1 (TNFAIP1) gene is a potential target of miR-15a and can be directly regulated by miR-15a. Down-regulation of TNFAIP1 induced effects on osteosarcoma cell lines similar to those induced by miR-15a. Taken together, these data suggest that miR-15a may act as a tumor suppressor, which is commonly down-regulated in both osteosarcoma tissues and cells. TNFAIP1 plays an important role in mediating miR-15a dependent biological functions in osteosarcoma. Reintroduction of miR-15a may be a novel therapeutic strategy by down-regulating TNFAIP1 expression.     

Laminin alpha1 chain in human renal cell carcinomas and integrin-mediated adhesion of renal cell carcinoma cells to human laminin isoforms

In human tissues, the laminin (Ln) alpha1 chain shows a restricted and developmentally regulated distribution in basement membranes (BMs) of a subset of epithelial tissues, including those of renal proximal convoluted tubules. The present study investigated the distribution of the Ln alpha1 chain in renal cell carcinomas (RCCs) and oncocytomas as well as in xenografted tumours induced in nude mice with four characterized RCC cell lines. These cell lines were also used in cell adhesion studies with purified laminins. By immunohistochemistry it was found that the Ln alpha1 chain is widely present in the BMs of RCCs, all of the specimens presenting immunoreactivity. High-grade RCCs tended to contain more BM-confined and stromal immunoreactivity than low-grade tumours, none of the grade 3 (G3) carcinomas being negative and all of the metastatic specimens showing partial or overall BM immunoreactivity. Double immunolabelling experiments showed that in RCC BMs but not in vessel walls, the Ln alpha1 chain was co-distributed with Ln alpha5, beta1, and beta2 chains, implying the presence of Ln-1/Ln-3 and Ln-10/Ln-11. In papillary RCCs, the Ln alpha1 chain co-localized with Ln-5. The oncocytomas lacked immunoreactivity for the Ln alpha1 chain. Xenografted tumours induced in nude mice showed BM-like deposition of the Ln alpha1 chain. In cell adhesion studies, mouse and human Ln-1 were equally effective in promoting cell adhesion of all RCC cell lines. For each cell line, Ln-10 and Ln-10/11 were equally effective adhesive substrates, all cell lines adhering more avidly to these laminins than to mouse or human Ln-1. As judged by inhibition assays employing specific integrin antibodies, adhesion of normal human renal proximal tubular epithelial (RPTE) cells and RCC cells from a G1 tumour to human Ln-1 was mediated mainly by alpha(6)beta(1) integrin, while only the G1 RCC cells adhered to mouse Ln-1 by using alpha(6)beta(1) integrin. For adhesion to Ln-10, RPTE cells and RCC cells from a G1 tumour used an unidentified beta(1) integrin. Cells from G3 tumours mainly used an alpha(3)beta(1) integrin complex for adhesion to mouse Ln-1 and to human Ln-1 and Ln-10. For all cells, adhesion to the Ln-10/11 mixture was mediated by an unidentified integrin complex or by other adhesion molecules. These results show that laminin trimers containing the alpha1 chain are, in contrast to oncocytomas, abundant in the BMs of RCCs. This is in keeping with their suggested origin from renal proximal tubular epithelium known for its capacity to produce the Ln alpha1 chain. The results also show that RCC cells utilize complex, mainly integrin alpha(3)beta(1)- and integrin alpha(6)beta(1)-mediated, mechanisms for adhesion to laminins. The adhesion to Ln-1 changes from integrin alpha(6)beta(1) to integrin alpha(3)beta(1) upon increasing malignancy and, especially for Ln-10 and Ln-10/11, other adhesion molecules of non-integrin type may contribute to the adhesion.     

VEGF165反义RNA对人食管鳞癌细胞影响的实验研究

目的 :观察血管内皮生长因子16 5 (VEGF16 5 )反义RNA对人食管鳞癌细胞EC10 9的影响 ,探讨其治疗食管癌的可行性。方法 :采用亚克隆技术 ,构建并鉴定VEGF16 5 反义RNA的真核表达载体。以重组质粒转染人食管鳞癌细胞EC10 9后 ,将其接种于裸鼠皮下 ,分别利用原位杂交、激光共聚焦、图象分析及微血管计数等方法 ,观察转染前后EC10 9细胞的生物学性状和致瘤性。结果 :成功地构建了VEGF16 5 反义RNA的真核表达载体 ,并在EC10 9细胞中获得表达。转染细胞中VEGF16 5 的表达下降 75% ,其生物学性状不受外源基因表达的影响 ,但其在裸鼠皮下的致瘤性和和瘤组织中血管的生成明显下降。VEGF16 5 反义RNA转染组、空载体转染组和对照组中肿瘤的体积 ,分别为 (82 0± 112 .5)mm3 、(793 0± 10 3 5)mm3 和 (7850± 950 )mm3(P <0 .0 1) ;微血管的密度分别为 (8.5± 1.2 ) /mm2 、(44.3± 9.4) /mm2 和 (46.4± 12 .6) /mm2 (P <0 .0 1)。结论 :VEGF16 5反义RNA能够明显减少食管鳞癌细胞内VEGF16 5 的表达 ,具有抑制肿瘤生长和血管生成的作用 ,可望用于实体肿瘤的辅助治疗。     

siRNA敲减缺氧诱导因子1α表达对口腔鳞癌细胞活力及癌胚抗原相关细胞黏附分子1表达的影响

目的:探究缺氧诱导因子1α(HIF-1α)与口腔鳞癌细胞活力和凋亡的关系以及作用机制。方法:采用RT-PCR和Western blot检测口腔鳞癌细胞系Tca8113和CAL27以及正常口腔上皮细胞NOK中HIF-1α和癌胚抗原相关细胞黏附分子1(CEACAM1)的mRNA和蛋白表达量; RNA干扰技术沉默口腔鳞癌CAL27细胞中HIF-1α的表达,实验分为空白对照组、无义对照组和siRNA-HIF1-α组,MTT实验检测敲减HIF-1α表达对细胞活力的影响,流式细胞术检测细胞凋亡率的变化,Western blot检测HIF-1α、P21、血管内皮生长因子(VEGF)、Bcl-2和Bax的蛋白水平。结果:HIF-1α和CEACAM1在口腔鳞癌细胞中的表达量显著高于正常口腔细胞(P 0. 05),且二者表达量呈正相关; HIF-1α和CEACAM1在CAL27细胞中的表达量显著高于Tca8113细胞(P 0. 05)。siRNA-HIF-1α组细胞中的HIF-1α蛋白表达量显著低于空白对照组(P 0. 05)。敲减HIF-1α表达显著抑制CAL27细胞的活力(P 0. 05),促进其凋亡(P 0. 05),显著增加P21和Bax的蛋白水平(P 0. 05),明显降低VEGF和Bcl-2的蛋白水平(P 0. 05)。结论:HIF-1α在口腔鳞癌中高表达,干扰HIF-1α的表达可显著抑制细胞的活力,促进其凋亡。这可能是通过调控HIF-1α下游靶基因的表达以及肿瘤血管的生成来发挥作用的。     

Effect of miR-200b on retinal endothelial cell function under high glucose environment

As one of the important complications of diabetes, diabetic retinopathy (DR) presented high incidence worldwide. Hyperglycemia is an important promoting factor for DR occurrence and development. It can damage retinal endothelial cell, resulting in retinal structure and function disorder. Studies have shown that miR-200b may involve in regulating DR occurrence and development, but its specific function and mechanism have not been elucidated. This study aimed to investigate miR-200b effect and mechanism on human retinal endothelial cells (hRECs) under high glucose environment. hRECs were cultured under high glucose or normal environment. Real time PCR was applied to detect miR-200b expression. MiR-200b was transfected to hRECs and MTT was used to detect its effect on hRECs proliferation under high glucose environment. Real time PCR and Western blot were performed to determine VEGF and TGFβ1 expression in the retina endothelial cells. MiR-200b expression decreased significantly under high glucose environment, whereas hRECs proliferated obviously. Compared with normal control, VEGF and TGFβ1 mRNA and protein expression increased markedly (P < 0.05). After miR-200b transfection, miR-200b expression increased, while VEGF and TGFβ1 mRNA and protein expression decreased obviously. Compared with high glucose group, hRECs proliferation was inhibited (P < 0.05). MiR-200b can regulate RECs growth and proliferation by changing VEGF and TGFβ1 expression to delay DR.     

人VEGF165基因真核表达质粒的构建及其对血管内皮细胞增殖的影响

目的 :构建人VEGF16 5基因的真核表达质粒pBudCE4 .1/VEGF16 5 ,观察其在血管内皮细胞 (VEC)中的表达和表达产物对VEC增殖的影响。方法 :用RT PCR法从引产胎儿心脏组织中克隆VEGF16 5基因 ,并将其克隆至真核表达质粒pBud CE4 .1中 ,对重组真核表达质粒pBudCE4 .1/VEGF16 5进行酶切鉴定和测序。以脂质体转染法将pBudCE4 .1/VEGF16 5导入VEC中 ,用Northernblot和免疫细胞化学染色法 ,分别从mR NA水平和蛋白质水平检测它们在转染的VEC中的表达 ,并检测表达产物对VEC增殖的影响。结果 :人VEGF16 5基因的RT PCR产物为 5 76bp。测序结果显示 ,扩增的VEGF16 5基因的序列与基因文库中登录的序列完全一致。经HindⅢ和BamHI酶切鉴定证实 ,VEGF16 5基因已成功地克隆至真核表达质粒pBudCE4 .1中。以其转染血管内皮细胞 (VEC)后 ,经Northernblot杂交和免疫细胞化学染色法检测均证实VEGF16 5基因表达。表达产物对VEC增殖有明显的促进作用。结论 :所构建的pBudCE4 .1/VEGF16 5真核表达质粒可在VEC中表达 ,表达产物可明显促进VEC增殖 ,为通过VEGF16 5基因转染防治移植器官内血管的狭窄奠定了基础     

Allelic loss on chromosomes 3p, 5q and 17p in renal cell carcinomas

Lossof-heterozygosity (LOH) has been studied on 3p (von Hippel-Lindau gene locus), 5q and 17p (p53 gene locus) by a polymerase chain reaction (PCR)-based strategy in 42 sporadic renal cell carcinomas (RCC). LOH at seven micro-satellite loci on 5q was Investigated because a tumor sup presser gene on 5q involved In the development and/or progression of RCC has not yet been identified. LOH was found In seven (17%) RCC at single or multiple locl on 5q, 38% (11/29 Informative cases) on 3p, and 6% (2/35 Informative cases) on 17p. Replication error (RER) was present in 10% (4/42) RCC at single or multiple loci. The minimum region of deletion on 5q to account for LOH was mapped to 5q31.1 (interferon regulatory factor-1; IRF-1 locus), where LOH was detected In 23% (6/26 Informative cases). LOH on 3p and 5q occurred In both stage 2 and more advanced (stage 3 and 4) tumors at similar incidences (41 and 33% on 3p; and 24 and 22% on 5q, respectively), suggesting that LOH on these chromosomes Is an early genetic event. All RCC exhibiting LOH on 3p or 5q (IRF-1 locus) were the clear cell or the mixed clear and granular cell types. These findings suggest that LOH on 3p and 5q plays an important role in the genesis of clear cell RCC. In addition, only one tumor exhibited LOH on both 3p and 5q, which suggests that LOH occurs not sequentially but independently.