Proliferating Cell Nuclear Antigen Labeling Research Articles

Patterns of cell proliferation and rod photoreceptor differentiation in shark retinas

We studied the pattern of cell proliferation and its relation with photoreceptor differentiation in the embryonic and postembryonic retina of two elasmobranchs, the lesser spotted dogfish ( Scyliorhinus canicula) and the brown shyshark ( Haploblepharus fuscus). Cell proliferation was studied with antibodies raised against proliferating cell nuclear antigen (PCNA) and phospho-histone-H3, and early photoreceptor differentiation with an antibody raised against rod opsin. As regards the spatiotemporal distribution of PCNA-immunoreactive cells, our results reveal a gradual loss of PCNA that coincides in a spatiotemporal sequence with the gradient of layer maturation. The presence of a peripheral growth zone containing pure-proliferating retinal progenitors (the ciliary marginal zone) in the adult retina matches with the general pattern observed in other groups of gnathostomous fishes. However, in the shark retina the generation of new cells is not restricted to the ciliary marginal zone but also occurs in retinal areas that contain differentiated cells: (1) in a transition zone that lies between the pure-proliferating ciliary marginal zone and the central (layered) retina; (2) in the differentiating central area up to prehatching embryos where large amounts of PCNA-positive cells were observed even in the inner and outer nuclear layers; (3) and in the retinal pigment epithelium of prehatching embryos. Rod opsin immunoreactivity was observed in both species when the outer plexiform layer begins to be recognized in the central retina and, as we previously observed in trout, coincided temporally with the weakening in PCNA labelling.

Inhibitory effect of bread crust antioxidant pronyl-lysine on two different categories of colonic premalignant lesions induced by 1,2-dimethylhydrazine

Colorectal malignancies continue to be one of the most frequent and life-threatening diseases throughout the world. Pronyl-lysine, a product obtained from bread crust, is a potent free radical scavenging antioxidant exerting chemopreventive activity by reducing oxidative stress. This study was conducted to investigate the effects of pronyl-lysine supplementation on the formation of colonic precancerous lesions, circulatory lipid peroxidation, and enzymic antioxidant status in 1,2-dimethylhydrazine-induced colon carcinogenesis. Male Wistar rats were randomized into seven groups; group 1 was control rats, group 2 received pronyl-lysine (2 mg/kg body weight orally) everyday, rats in groups 3-7 were administered subcutaneous 1,2-dimethylhydrazine (20 mg/kg body weight) once a week for 15 consecutive weeks. In addition, group 4 (pre-initiation), 5 (initiation), 6 (post-initiation), and 7 (entire period) received pronyl-lysine (2 mg/kg body weight orally) everyday. At the end of 34 weeks, indicative markers of lipid peroxidation and changes in antioxidant defense system were measured in circulation. The results showed that 1,2-dimethylhydrazine significantly increased total aberrant crypt foci formation, total number of dysplastic foci, beta-catenin accumulated crypts and proliferating cell nuclear antigen labeling index in the colon, and enhanced lipid peroxidation markers and decreased enzymic antioxidant activities in the plasma and erythrocyte lysate as compared with untreated controls. Pronyl-lysine supplementation significantly reversed the changes as compared with the rats treated with 1,2-dimethylhydrazine alone. The effect of pronyl-lysine was more pronounced when supplemented throughout the study period (group 7). These findings suggest that pronyl-lysine suppresses 1,2-dimethylhydrazine-induced colon carcinogenesis effectively.

Low‐dose simultaneous delivery of adenovirus encoding hepatocyte growth factor and vascular endothelial growth factor in dogs enhances liver proliferation without systemic growth factor elevation

Hepatocyte growth factor (HGF) and vascular endothelial growth factor (VEGF) gene transfer proved to enhance liver regeneration. However, elevation of their plasma levels may induce potentially serious distant effects such as tumorigenesis or proliferative retinopathy. This study was performed to examine whether simultaneous administration of low-dose adenovirus encoding HGF and VEGF genes in dogs will stimulate liver proliferation but without inducing liver toxicity or systemic elevation of HGF and VEGF levels. Adult dogs received an intravenous injection of low-dose adenoviral vectors encoding human HGF and VEGF (HGF/VEGF), beta-galactosidase (lacZ) or phosphate-buffered saline (PBS). Liver proliferation was measured using the proliferating cell nuclear antigen (PCNA) immunostaining labelling index. HGF and VEGF plasma concentrations and transaminases were repeatedly measured. Transgene expression was evaluated using reverse-transcription polymerase chain reaction. Human HGF and VEGF expressions were detected only in the liver of HGF/VEGF dogs at day 2 after injection but declined at sacrifice (day 7). No expression was detected in the liver of the lacZ or PBS groups. Plasma levels of HGF and VEGF were not statistically different from those in the lacZ group (P=0.81, P=0.22 respectively). The PCNA labelling index was five-fold higher in the HGF/VEGF group compared with the lacZ group (P<0.01). No immunostaining was detected in the PBS group. Transaminases were only elevated (P<0.01) in the lacZ group compared with the other groups. We showed that simultaneous administration of low-dose adenoviral vectors encoding human HGF and VEGF genes can induce transgene expression and liver proliferation without liver toxicity or systemic growth factor elevation.

The HP1α–CAF1–SetDB1‐containing complex provides H3K9me1 for Suv39‐mediated K9me3 in pericentric heterochromatin

Trimethylation of lysine 9 in histone H3 (H3K9me3) enrichment is a characteristic of pericentric heterochromatin. The hypothesis of a stepwise mechanism to establish and maintain this mark during DNA replication suggests that newly synthesized histone H3 goes through an intermediate methylation state to become a substrate for the histone methyltransferase Suppressor of variegation 39 (Suv39H1/H2). How this intermediate methylation state is achieved and how it is targeted to the correct place at the right time is not yet known. Here, we show that the histone H3K9 methyltransferase SetDB1 associates with the specific heterochromatin protein 1alpha (HP1alpha)-chromatin assembly factor 1 (CAF1) chaperone complex. This complex monomethylates K9 on non-nucleosomal histone H3. Therefore, the heterochromatic HP1alpha-CAF1-SetDB1 complex probably provides H3K9me1 for subsequent trimethylation by Suv39H1/H2 in pericentric regions. The connection of CAF1 with DNA replication, HP1alpha with heterochromatin formation and SetDB1 for H3K9me1 suggests a highly coordinated mechanism to ensure the propagation of H3K9me3 in pericentric heterochromatin during DNA replication.

Diffusion‐weighted imaging of prostate cancer: Correlation between apparent diffusion coefficient values and tumor proliferation

To investigate whether the apparent diffusion coefficient (ADC) values of prostate cancer (PCa) are able to reflect tumor proliferation. The clinical and pathological information for 38 patients with PCa and 33 patients with benign prostate hyperplasia (BPH) were studied. Examination of the patients was performed using a 1.5 T superconducting magnetic scanner equipped with a pelvic phased-array multicoil. Diffusion-weighted images (DWIs) were acquired using an echo-planar imaging sequence. The ADC values of PCa, BPH, and peripheral zone (PZ) were calculated. The cellularity of PCa was recorded based on hematoxylin and eosin staining. The proliferating cell nuclear antigen (PCNA) was detected using an immunohistochemical technique. The ADC values of PCa, BPH, and PZ were 49.32 +/- 12.68 x 10(-5) mm(2)/s, 86.73 +/- 26.75 x 10(-5) mm(2)/s, and 126.25 +/- 27.21 x 10(-5) mm(2)/s, respectively. The ADC values of PCa were lower than those of BPH and PZ (P < 0.05). The cellularity and PCNA labeling index (LI) of PCa were higher than those of BPH (P < 0.05). The ADC values of PCa were negatively correlated with those of cellularity and PCNA LI (r = -0.646 and -0.446, respectively; P < 0.05). The ADC values of PCa can reveal the differences in proliferative activity between PCa and BPH. These values are therefore able to predict the proliferative rate of variously differentiated prostate cancers.

Beneficial effects of splenectomy on massive hepatectomy model in rats

Possible spleno-hepatic relationships during hepatectomy remain unclear. The purpose of this study was to investigate the impact of splenectomy during massive hepatectomy in rats. Rats were divided into the following two groups: 90% hepatectomy (Hx group), hepatectomy with splenectomy (Hx+Sp group). The following parameters were evaluated; survival rate, biochemical parameters, quantitative RT-PCR for hemeoxygenase-1 (HO-1) and tumor necrosing factor alpha (TNFalpha), immunohistochemical staining for HO-1, proliferating cell nuclear antigen labeling index and liver weights. The survival rate after massive hepatectomy significantly improved in Hx+Sp group as well as serum biochemical parameters, compared with Hx group (P < 0.05). HO-1 positive hepatocytes and its mRNA expression significantly increased and TNFalpha mRNA expression significantly decreased in Hx+Sp group compared with Hx group (P < 0.05). Moreover, liver regeneration was significantly accelerated at 48 and 72 h after hepatectomy in Hx+Sp group. Splenectomy had beneficial effects on massive hepatectomy by ameliorating liver injuries and promoting preferable liver regeneration.

Regeneration of Graft Livers and Limited Contribution of Extrahepatic Cells After Partial Liver Transplantation in Humans

Background Liver regeneration is still not fully understood. Partial liver transplantation (LT) can provide the opportunity to investigate the mechanisms of liver regeneration, including the contribution of extrahepatic cells to liver regeneration. Methods Of 61 patients transplanted with partial liver graft between August 1997 and October 2006, 56 patients were studied, including 49 adults and 7 children. Sequential computed tomography volumetric analysis was performed for volume measurement, while proliferating cell nuclear antigen (PCNA) labeling index was investigated for liver cell proliferation in nonprotocol liver biopsy specimens. In addition, 15 male recipients who had female liver grafts were investigated in order to detect Y chromosomes as extrahepatic cells in nonprotocol liver biopsy specimens. Results Graft volume per standard liver volume was markedly increased after adult-to-adult living-donor (LD) LT. In pediatric transplants, there was no volume increase over time. PCNA labeling index was vigorous in adult-to-adult LDLT in the early period after LDLT. No Y chromosome was evident in hepatocytes from female-donor male-recipient grafts during or after liver regeneration. However, in the cases of failing grafts of this type, many Y-chromosome-positive cells were observed in the graft liver. The character of those cells was CD34(−), CK9(−), hepatocyte-specific antigen(−), and CD68(+/−). Conclusion In adult-to-adult LDLT, vigorous liver regeneration occurs in the graft liver, demonstrated by not only volumetric but cell kinetic analysis. Involvement of extrahepatic cells in normal liver regeneration seems limited.

Two-Stage Portal Vein Ligation Facilitates Liver Regeneration in Rats

Background/Aims: Recent reports have demonstrated that some patients are unable to undergo scheduled liver resection after preoperative portal vein embolization due to insufficient hypertrophy of the future remnant liver. The present study examined whether two-stage portal vein ligation (PVL) increases hypertrophy of the future remnant liver compared to conventional PVL in rats. Methods: Rats were divided into 3 groups: group A, ligation of left primary branch; group B, ligation of right and left primary branches; group C, ligation of the left primary branch, followed by 2-stage PVL 7 days postoperatively. To evaluate liver regeneration, the proliferating cell nuclear antigen labeling index (LI), mitotic index (MI) in the caudate lobe and weight ratio of caudate lobe to body weight were measured. Results: The weight ratio of caudate lobe to body weight was significantly higher in group C than in groups A or B 14 days postoperatively. In groups A and B, LI and MI in the caudate lobe peaked 2 days postoperatively, then decreased to preoperative levels by 7–8 days postoperatively, but remained significantly elevated until 10–14 days postoperatively in group C. Conclusion: Two-stage PVL increases hypertrophy of the future remnant liver compared to conventional PVL in rats.

Effects of Bicyclol on Liver Regeneration After Partial Hepatectomy in Rats

Bicyclol is a synthetic antihepatitis drug with antioxidative property. The present study was performed to investigate the effect of bicyclol on liver regeneration after partial hepatectomy in rats. Bicyclol (300 mg/kg) was given to rats subjected to 70% hepatectomy three times before operation. At 6, 24, and 48 h after resection, samples were collected for the measurement of serum alanine aminotransferase (ALT), total bilirubin (TBil), hepatic glycogen, malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH). Moreover, liver regeneration rate, proliferating cell nuclear antigen (PCNA) labeling, proliferation index, and histopathological examination were evaluated at 48 h after hepatectomy. As a result, bicyclol significantly increased regeneration rate, mitotic index (MI), PCNA labeling index, and proliferation index in PH rats. Additionally, bicyclol remarkably inhibited the elevation of serum ALT and TBil levels, alleviated the formation of liver MDA, restored impaired antioxidant SOD and GSH, increased hepatic glycogen content, and also attenuated hepatic vacuolar degeneration. These results suggested that bicyclol had a beneficial effect on liver regenerative capacity of the remnant liver tissue after hepatectomy, probably due to its antioxidative property.

Regression of flow-induced pulmonary arterial vasculopathy after flow correction in piglets

Chronic thromboembolic pulmonary hypertension is due to partial obstruction of the pulmonary arterial bed and may resolve after pulmonary thromboendarterectomy. Persistent pulmonary hypertension, the main complication after pulmonary thromboendarterectomy, may reflect vessel alterations induced by high flow in unobstructed lung territories. The aim of this study was to determine whether correcting high flow led to reversal of the vasculopathy in piglets. The effects of high pulmonary blood flow were investigated 5 weeks after creation of an aortopulmonary shunt (n = 10), and reversibility of vessel disease was evaluated at 1 week (n = 10) and 5 weeks after shunt closure (n = 10), compared to sham-operated animals (n = 10). Hemodynamic variables, pulmonary artery reactivity, and morphometry were recorded. We also investigated the endothelin, angiopoietin, and nitric oxide synthase pathways. High flow increased medial thickness in distal pulmonary arteries (55.6% +/- 1.2% vs 35.9% +/- 0.8%; P < .0001) owing to an increase of smooth muscle cell proliferation (proliferating cell nuclear antigen labeling). The endothelium-dependent relaxation was altered (P < .05). This phenomenon was associated to an overexpression of endothelin-1, endothelin-A, angiopoietin 1, angiopoietin 2, and Tie-2 (P < .05). After 1 week of shunt closure, all overexpressed genes returned to control values, the proliferation of smooth muscle cells stopped, and smooth muscle cell apoptosis increased (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling), preceding the normalization of the wall thickness hypertrophy and the pulmonary artery vasoreactivity observed at 5 weeks after shunt closure. These results demonstrate that endothelin-1 and angiopoietin pathways are involved in vasculopathy development and may be important therapeutic targets for preventing persistent pulmonary hypertension after pulmonary thromboendarterectomy.

Long-term effect of Spirulina platensis extract on DMBA-induced hamster buccal pouch carcinogenesis (immunohistochemical study)

In cancer research, the use of complementary and alternative medicine has increased over the past decade. In this study, 80 male golden Syrian hamsters were divided into four equal groups; the right buccal pouches of the hamster rats in group 1 were painted with 0.5% solution of 7, 12-dimethylbenz[a]anthracene (DMBA), three times a week for 32 weeks. The same pouches of group 2 were subjected to the same DMBA painting; but at the same time, the animals received 10 mg/daily Spirulina platensis extract for the same period. In group 3, the same regimen of DMBA painting was done but for 24 weeks only and the daily systemically S. platensis was received for the 32 weeks. In group 4, neither DMBA painting nor S. platensis administration was done but pouches were painted with saline and served as a control one. Five rats from each group were sacrificed at 12, 24, 28, and 32 weeks, respectively. The required pouches were excised, fixed, and embedded in paraffin to be immunostained with proliferating cell nuclear antigen (PCNA). The results showed that increased PCNA expression was directly related to the severity of pathological alterations from normal epithelium to dysplasia and from dysplasia to squamous cell carcinoma (SCC) in the study groups at the different extended periods of DMBA application and S. platensis extract administration. Analysis of variance and Duncan's multiple-range test for PCNA labeling index were proved a high significant difference (P < 0.01) between the different groups. From the previous results, it can be concluded that S. platensis extract has a beneficial role in regression of cancer progression.

Suppression of Right Ventricular Hypertrophy After Extensive Pulmonary Resection in Rats by Granulocyte Colony-Stimulating Factor

The objective of the present study was to investigate the effects of granulocyte colony-stimulating factor (G-CSF) on right ventricular hypertrophy following extensive pulmonary resection in rats. Adult rats were divided into four groups: (1) Group S (right thoracotomy only); (2) Group L (right three lobectomy); (3) Group LG10 (Group L+G-CSF [10microg/kg/d]); and (4) Group LG100 (Group L+G-CSF [100microg/kg/d]). At postoperative day 21, weight ratio of the right ventricular to the left ventricle plus septum (RV/LV+S, indicator of right ventricular hypertrophy) were measured, and a histopathological study was conducted to determine percentage wall thickness of peripheral pulmonary arteries and proliferating cell nuclear antigen labeling index (indicator of oxidative DNA damage) of right ventricles. Mean RV/LV+S for Group S was 0.27+/-0.02, significantly smaller than that for the lobectomy groups (Group L, LG10, LG100; 0.47+/-0.05, 0.35+/-0.02, 0.38+/-0.05). G-CSF significantly suppressed right ventricular hypertrophy. Mean medial wall thickness of peripheral pulmonary arteries for Group S was 13.6% +/- 4.9%, significantly smaller than that for Group L (22.9% +/- 9.6%). Compared with Group L, G-CSF reduced medial wall thickness (LG10, 17.6% +/- 9.5%; LG100, 18.0% +/- 11.2%). Incidence of proliferating cell nuclear antigen positive nuclei for Group S was 1.07% +/- 0.49%, significantly smaller than that for Group L (13.77% +/- 5.87%). G-CSF significantly reduced the incidence of proliferating cell nuclear antigen positive nuclei (LG10, 4.04% +/- 2.14%; LG100, 3.18% +/- 1.66%). G-CSF administration not only reduce medial wall thickness of peripheral pulmonary arteries but also directly protect cardiomyocytes of the right ventricle, thus suppressing right ventricular hypertrophy. These results suggest that low-dose G-CSF administration can prevent right heart failure following extensive pulmonary resection.

Platelets contribute to the reduction of liver fibrosis in mice

Several recent studies have reported that liver cirrhosis (LC) can be ameliorated, but few adequate strategies are available against liver fibrosis. Although LC clinically shows thrombocytopenia and hypersplenism, the correlation with liver fibrosis and platelets remains unclear. The aim of the present study was to investigate the effect of platelets on liver fibrosis in mouse models. To induce liver fibrosis, C57BL6 female mice were injected i.p. with 1 mL/kg carbon tetrachloride (CCl(4)) twice a week for 8 weeks. Thrombocytosis was achieved by giving thrombopoietin or splenectomy in addition to CCl(4) intoxication. At 8 weeks, whole blood and liver specimens were obtained for studies as follows: peripheral platelet counts, histopathological examination, hydroxyproline assay, immunostaining, quantification of mRNA expression, and microarray analysis. Thrombocytosis significantly reduced liver fibrosis and hydroxyproline content of liver tissues compared to mice with CCl(4) administration alone. Platelets suppressed increments in mRNA expression for transforming growth factor-beta, and increased matrix metalloproteinase-9 expression in the liver. Microarray analysis of the liver revealed that platelets upregulated gene expressions involved in cell proliferation compared to expression in mice with CCl(4) intoxication alone. Platelets also increased liver volume, proliferative cell nuclear antigen labeling index, and mitotic index in fibrotic mice. These results clearly show that platelets reduce liver fibrosis and promote liver regeneration, even under cirrhotic conditions. We, therefore, propose that platelets could offer a potent tool in the treatment of liver cirrhosis.

The Value of Survivin Gene and Proliferation of Hepatocytes in Screening for Hepatocellular Carcinoma.

BackgroundThe prospective surveillance programs on patients with liver diseases based on repeat ultrasound examinations of liver and serum α-fetoprotein (AFP) detection were reported having the probability of finding hepatocellular carcinoma (HCC) at its early stage, but it is time-consuming and not cost-effective. To improve the effectiveness and cost-effective of HCC surveillance program, close monitoring has been focused on patients with liver cirrhosis who have particularly high risk of HCC development. It has been found that high liver cell proliferation is a reliable predictor of HCC development, and survivin is a new gene with the role of suppressing apoptosis which has been studied mostly over the past few years. This study aimed to evaluate the usefulness of proliferation cell nuclear antigen (PCNA) and survivin detection in the process of screening cirrhotic patients with high risk of HCC development.MethodsTotal RNA was extracted from fresh specimens of HCC and liver cirrhosis. Survivin mRNA amplification was performed by reverse transcription polymerase chain reaction (RT-PCR). Immunostaining for PCNA was employed to assess liver cell proliferation activity in formalin-fixed, paraffin embedded liver specimens. Five liver specimens obtained from patients operated for hemangioma were used as normal control. The PCNA labeling index was determined as the mean value of positive cells in ten different microscopic fields.ResultsRT-PCR was performed in 17 HCC and 10 liver cirrhosis specimens, 11 HCC specimens showed 344bps molecular survivin DNA band in 1% agarose electrophoresis, but none of liver cirrhosis specimens showed positive band. The survivin positive rate in HCC specimens was 64.7% (11/17); The PCNA labeling index was 2.38 ± 2.11 in 30 liver cirrhosis specimens, while 10.08 ± 12.28 in 30 HCC specimens, the latter was significantly higher than the former, P = 0.003. The median PCNA labeling index of 11 survivin positive HCC specimens was 6.8 (from 0.5 to 40), which is significantly higher than that of 6 survivin negative HCC specimens (2.15).ConclusionsSurvivin expressed in HCC tissues but not in liver cirrhosis tissues, this phenomenon indicates that the gene expression may occur at the late phase of liver cell cancer transformation. Compared with survivin detection, the PCNA detection on liver cells of cirrhosis patients is better to differentiate high-risk HCC transformation among liver cirrhosis patients.

Toll-Like Receptor 3 Is a Negative Regulator of Embryonic Neural Progenitor Cell Proliferation

Toll-like receptors (TLRs) play important roles in innate immunity. Several TLR family members have recently been shown to be expressed by neurons and glial cells in the adult brain, and may mediate responses of these cells to injury and infection. To address the possibility that TLRs play a functional role in development of the nervous system, we analyzed the expression of TLRs during different stages of mouse brain development and assessed the role of TLRs in cell proliferation. TLR3 protein is present in brain cells in early embryonic stages of development, and in cultured neural stem/progenitor cells (NPC). NPC from TLR3-deficient embryos formed greater numbers of neurospheres compared with neurospheres from wild-type embryos. Numbers of proliferating cells, as assessed by phospho histone H3 and proliferating cell nuclear antigen labeling, were also increased in the developing cortex of TLR3-deficient mice compared with wild-type mice in vivo. Treatment of cultured embryonic cortical neurospheres with a TLR3 ligand (polyIC) significantly reduced proliferating (BrdU-labeled) cells and neurosphere formation in wild type but not TLR3(-/-)-derived NPCs. Our findings reveal a novel role for TLR3 in the negative regulation of NPC proliferation in the developing brain.

Metalloproteinases 2 and -9 activity during promotion and progression stages of rat liver carcinogenesis

Activity of metalloproteinases 2 and 9 (MMP-2 and 9) during promotion and progression of rat liver carcinogenesis was investigated in a modified resistant hepatocyte model. Development of preneoplastic liver lesions positive for glutathione S-transferase 7-7-(GST-P 7-7-positive PNL) and tumors besides hepatocytes positive for proliferating cell nuclear antigen (PCNA) were quantified and compared to MMP-2 and-9 activity using gelatin zymography. Marked increases in GST-P 7-7-positive PNL development, PCNA labeling indices, MMP-2 (pro, intermediate and active forms) and pro-MMP-9 activity were observed after proliferative stimulus induced by 2-acetylaminofluorene (2-AAF) exposure cycles. After 2-AAF withdrawal, increase in MMP-2 activity was detected only in neoplastic mixed lesions, whereas active MMP-9 was increased in both PLN and neoplastic tissues. Our findings suggest that MMP-2 may be associated with proliferative events induced by 2-AAF rather than with selective growth of PNL and that MMP-9 could be associated with progression of PNL and neoplastic mixed lesions.

Dynamic contrast-enhanced magnetic resonance imaging for estimating tumor proliferation and microvessel density of oral squamous cell carcinomas

We evaluated the relationship between histopathological prognostic factors, tumor proliferation microvessel density (MVD), and enhancement parameters in dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in oral squamous cell carcinoma (SCC). Twenty-eight T2 and T3 patients with primary oral SCC underwent DCE-MRI using three-dimensional fast imaging with a steady-state precession sequence. Tumor cell proliferation and MVD of all surgical specimens were evaluated using immunohistochemical staining with CD34 and the antibody for proliferating cell nuclear antigen (PCNA). Regression analysis was used to statistically analyze the relationship between the PCNA labeling index or MVD and each of three DCE-MRI parameters: maximum CI (CI-max), maximum CI gain (CI-gain) and the CI-gain / CI-max ratio). The PCNA labeling index and MVD showed significant correlations with the CI-gain/CI-max ratio (P=0.0012, r=0.581 and P=0.00141, r=0.574, respectively). The assessment of DCE-MRI parameters may prove to be a valuable non-invasive method for assessing tumor cell proliferation and MVD of patients with oral cancer.

Development of the human insular cortex: differentiation, proliferation, cell death, and appearance of 5HT-2A receptors

The development of the human insula was studied in the foetuses from 21 to 32 gestation weeks, using silver staining, immunohistochemistry of proliferative cell nuclear antigen (PCNA), activated caspase-3, and TdT-mediated dUTP-biotin nick end labeling (TUNEL) techniques. To test whether the insula also has a significant role in psychiatry behavior, we also mapped the major receptor of serotonin, 5HT-2A, in the developing insula as well. Opercular formation was evident by 21 weeks gestation. At this time, the cortical layers in the insula had started to organize, with silver impregnated pyramidal and stellate cells demonstrated various processes. By 25 gestation weeks, gyri in the insula were observed. PCNA positive cells decreased in density from 21 gestation weeks onwards while an increase of TUNEL positive cells was evident from 25 to 32 weeks of gestation. Activated capase-3 positive cells were detected in the insula, along with TUNEL positive cells, confirming possibly apoptosis. Serotonin 2A receptor appeared robustly in the 31/32 gestation week specimens. Our study showed early differentiation in the insula, when compared with other parts of the human cortex in the literature. In addition, proliferation as well as apoptosis were demonstration. Expression of 5HT (serotonin) 2A receptor positive cells in development was indicative of the insula as a significant psychiatric center.

Predicting lymph node status in early gastric cancer

Accurate prediction of lymph node (LN) status is of crucial importance for appropriate treatment planning in patients with early gastric cancer (EGC). However, there is no definitive consensus yet on which patient and/or tumor characteristics are associated with LN metastasis. A systematic search for studies investigating the relationship between patient and/or tumor characteristics and LN metastasis in EGC was performed in PubMed/MEDLINE. Patient and/or tumor characteristics associated with LN metastasis were identified by meta-analyzing results of individual studies. Forty-five studies were included. Variables significantly associated with LN metastasis in gastric cancer limited to the mucosa were: age younger than 57 years, tumor location in the middle part of the stomach, larger tumor size, macroscopically depressed tumor type, tumor ulcerations, undifferentiated tumors, diffuse tumor type according to the Lauren classification, lymphatic tumor invasion, tumors with a proliferating cell nuclear antigen (PCNA) labeling index of more than 25%, and matrix metalloproteinase-9-positive tumors. Variables significantly associated with LN metastasis in gastric cancer limited to the submucosa were: female sex, tumor location in the lower part of the stomach, larger tumor size, undifferentiated tumors, increasing depth of submucosal invasion, lymphatic tumor invasion, vascular tumor invasion, increased submucosal vascularity, tumors with a PCNA labeling index of more than 25%, tumors with a gastric mucin phenotype, and vascular endothelial growth factor-C-positive tumors. We identified several variables associated with LN metastasis in EGC. These variables should be included in future research, in order to assess which of these variables remain as significant predictors of LN metastasis.

Hypoxia-inducible factor 1 alpha in oral squamous cell carcinoma and its relation to prognosis

The aim of this study was to investigate the correlation between the expression of hypoxia-inducible factor 1 alpha (HIF-1 alpha) and proliferative activity in tumor cells, lymph node metastasis, as well as prognosis in patients with oral squamous cell carcinoma (OSCC). Fifty-seven biopsy specimens of OSCC were investigated for the expression of HIF-1 alpha and proliferating cell nuclear antigen (PCNA) by immunohistochemistry. None of the patients had received any prior treatments. The percentage of HIF-1 alpha immunopositive area (PHIA) was calculated using computer-assisted image analysis for quantitative assessment of HIF-1 alpha expression. The PCNA labeling index (LI) was evaluated as a proliferation marker. We found that the mean PHIA in all stages was 12.1% in the poor prognosis patients, and it was 6.4% in the good prognosis patients. There was a significant difference of PHIA between poor prognosis and good prognosis patients (P=0.0065). Furthermore, the mean PHIA in the patients who had no metastatic lymph nodes was 7.5%, while it was 11.7% in the patients who had metastatic lymph nodes. There was also a significant difference of PHIA between patients who had no metastatic lymph nodes and those who had metastatic lymph nodes (P=0.0487). On the other hand, significant correlation between PHIA and PCNA LI was not observed. These results provide the clinical data indicating that HIF-1 alpha may play an important role in lymph node metastasis and prognosis in patients with OSCC.