Proliferating Cell Nuclear Antigen Counts Research Articles

Nanocurcumin versus mesenchymal stem cells in ameliorating the deleterious effects in the cadmium‐induced testicular injury: A crosstalk between oxidative and apoptotic markers

Cadmium (Cd), a grave occupational pollutant, can result in; testicular damage. This study was designed to distinguish the potential effect of bone marrow-derived mesenchymal stem cells (BM-MSCs) versus that of curcumin nanoemulsion on Cd-induced testicular damage. Fifty adult male Sprague Dawley rats were distributed into five groups; control, sham control, Cd-treated, stem cell-treated and nanocurcumin-treated groups. Histological, immune histochemical; caspase 3 and proliferating cell nuclear antigen (PCNA) and CD 68, testosterone levels, nitric oxide, malondialdehyde (MDA)/glutathione (GSH) superoxide, dismutase (SOD), Western blot; B-cell lymphoma (Bcl-2), BCL2-Associated X Protein (BAX), BAX/Bcl-2 ratio and morphometry were done. Cadmium-treated group showed degenerated, detached seminiferous tubules, vacuolations and wide interstitial spaces containing fluid exudates. The same group revealed increased expression of BAX, BAX/Bcl-2 ratio, caspase 3, CD 68 and increased mean values of MDA, NO. Concomitantly, Cd has significant reduction in PCNA, Bcl-2 and sperm cell count when compared to control group. BM-MSCs- and nanocurcumin-treated groups revealed well-structured tubules and were perceived to expressively enhance the deleterious changes induced by Cd. The injurious changes on the testis induced by Cd were obviously improved when treated with either MSCs or nano-curcumin. BM-MSCs exerted more ameliorative changes.

Does ischemia-reperfusion injury affect ovarian reserve and follicle viability in a rat model with adnexal torsion?

Objective(s)To evaluate the effect of ischemia-reperfusion on follicle count and viability of follicles in a rat model with adnexal torsion. Study designA total of 24 female adult Wistar albino rats were included in the study. In the first laparotomy, right ovaries were twisted at 720° in the counterclockwise for a duration of two (n=12), four (n=6) or 16hours (n=6), whereas contralateral sites were kept as controls. In the second laparotomy, the right ovaries were detorsioned. Twenty-eight days after the detorsion, both ovaries were retrieved with the third laparotomy and placed in formaldehyde for subsequent examination under light microscopy and immunohistochemistry for proliferating cell nuclear antigen (PCNA). Besides histological evaluation by only using a standard scoring system, the viability of the follicles was also assessed by PCNA with immunohistochemical staining. ResultsMacroscopically, after two, four and 16hours of torsion, all the twisted ovaries had enlarged and demonstrated a dark, dusky appearance. However, under light microscopy with 10X magnification, the mean primordial+primary follicle count was comparable in twisted and control sides (26.7±7.6 vs. 28.1±5.9 respectively). In terms of the means of total tissue damage, only the loss of cohesion was found to be significantly different compared to the control sides (2.78±0.86 vs. 2.17±0.86, p=0.017). Immunohistochemical staining revealed significantly lower PCNA counts in the 16-hour torsion group only. Conclusion(s)Even though ovarian reserves, reflected by primordial and primary follicle count, are similar between torsioned and contralateral ovaries, the viability of the remaining follicles might be affected by a long-standing ischemia-reperfusion phenomenon. Follicle count may reflect an idea of fertility potential, but immunohistochemical staining using viability markers such as PCNA confirms the viability of the counted follicles and appears to be a more precise approach necessary for demonstrating the functional status.

Targeting pericytes with a PDGF-B aptamer in human ovarian carcinoma models

Purpose: On the basis of the known role of platelet-derived growth factor (PDGF)-BB/PDGF receptor (PDGFR) β in pericyte regulation, highly specific inhibitors of this target are needed. We tested the efficacy of a highly selective aptamer against PDGF-B with or without anti-VEGF therapy in ovarian cancer models. Experimental Design: The therapeutic efficacy of targeting endothelial cells (bevacizumab) and/or pericytes (PDGF-aptamer, AX102) was examined using HeyA8 and SKOV3ip1 orthotopic models of ovarian cancer metastasis. Following therapy, tumors were examined for microvessel density (MVD), proliferating cell nuclear antigen (PCNA), and vascular maturation (pericyte coverage). Results: Bevacizumab inhibited tumor growth by 45% and 48% in the HeyA8 and SKOV3ip1 models, respectively. AX102 had minimal effect on the HeyA8 model, but increased tumor growth in the SKOV3ip1 model. However, bevacizumab plus AX102 was more effective than bevacizumab alone, and resulted in 76 - 88% inhibition of tumor growth in both models. A longitudinal study in the HeyA8 model using bioluminescence imaging showed that combination of bevacizumab, AX102 and paclitaxel caused tumor reduction by 65% (based on bioluminescence imaging). In the HeyA8 model, MVD and PCNA counts were significantly reduced in the bevacizumab treatment groups, and pericyte coverage was significantly decreased in the AX102 treatment groups. In the SKOV3ip1 model, MVD and PCNA was significantly reduced in the bevacizumab treatment group, and even lower in the bevacizumab and AX102 combination treatment group. Conclusions: Dual targeting of endothelial cells and pericytes holds potential as an anti-vascular therapeutic approach in ovarian carcinoma.

Retrospective – systematic study and quantitative analysis of cellular proliferation and apoptosis in normal, hyperplastic and neoplastic perianal glands in dogs

Neoplasms in the perianal region are frequently diagnosed in dogs. The aetiology is unknown, and most of them are benign. In this study, 240 neoplasms of the perianal glands of dogs were retrieved from the Department of Pathology archives of the Faculty of Veterinary Medicine and Zootechny of University of São Paulo (FMVZ/USP), from 1984 to 2004. All 240 cases were re-examined by two pathologists. Nine cases (4%) were diagnosed as hyperplasia, 49 (20%) as group I adenoma, 81 (34%) were classified as moderately differentiated adenomas of the group II, 46 (19%) were poorly differentiated adenomas of group II, 48 (20%) were carcinoma of the group III according to the classification proposed by Berrocal, and 7 (13%) were other kind of tumours. Males over 8 years of age were predominantly affected. Cell proliferation was quantified by counting proliferating cell nuclear antigen (PCNA) positive nuclei, and apoptosis was quantified by counting fluorescent eosin-stained apoptotic corpuscles (AC) in normal tissue, hyperplasia and in different histologic types of neoplasia of these glands. A parallel pattern of increase in both parameters (cell proliferation and apoptosis) was obtained. The net growth index (NGI), represents how much a cell population is proliferating or dying and was achieved by dividing the mean PCNA count in 1000 cells by the mean AC stain count in 1000 cells. NGI was different between hyperplasia and neoplasia; group I adenomas have a much higher potential of growth, and NGI decreases from benign towards malignant lesions. These results show up the importance of studying cell proliferation and apoptosis to understand the carcinogenesis of dog perianal gland.

Orally administered penicillamine is a potent inhibitor of neointimal and medial thickening in porcine saphenous vein–carotid artery interposition grafts

In patients who have undergone coronary artery bypass grafting, blood copper levels are elevated for 6 weeks after surgery. Copper is an established risk factor for cardiovascular disease and atherogenesis and promotes oxidative stress, lipid oxidation, cell proliferation, and matrix formation, all components of vein graft disease. This project therefore examined the effect of the copper chelator penicillamine on saphenous vein graft thickening in a pig model. Saphenous vein-carotid artery interposition grafts were carried out in Landrace pigs. Penicillamine (10 mg/kg once daily, n = 8) was administered orally incorporated into small amounts of mashed potato for 1 month (n = 8 controls). Vein grafts were then excised and fixed at 100 mm Hg, histologic sections were prepared, and morphometry and measurement of proliferating cell nuclear antigen count were carried out. In vitro studies on the effect of copper or penicillamine on human vascular smooth muscle cell replication was carried out with bromodeoxyuridine incorporation. Administration of penicillamine had a potent inhibitory effect on both neointimal and medial thickness and proliferating cell nuclear antigen count but elicited a marked increase in luminal area and reduced serum copper concentrations. In vitro, copper augmented vascular smooth muscle cell proliferation, an effect blocked by penicillamine. Penicillamine alone also inhibited in vitro vascular smooth muscle cell replication. The administration of penicillamine reduces vein graft thickening and promotes positive remodeling through negation of copper-induced cell proliferation. Copper chelators may therefore be therapeutically useful in preventing late vein graft failure in patients undergoing reconstructive arterial surgery.

Immunoexpression of hepatocyte growth factor and c-Met receptor in the eutopic endometrium predicts the activity of ectopic endometrium

Objective To investigate the mitogenic and angiogenic activity of the eutopic and ectopic endometrium throughout the menstrual cycle and to examine whether the activity of the eutopic endometrium is useful to predict greater activity of the ectopic endometrium. Design Controlled clinicopathologic study using intact tissue. Setting Nagasaki University School of Medicine, Nagasaki, Japan. Patient(s) Fifteen infertile women with pelvic endometriosis and 10 women without endometriosis undergoing laparoscopy. Intervention(s) Biopsies from the ectopic endometrium and the corresponding eutopic endometrium were collected. Immunohistochemical staining was performed using respective antibodies, and a computer analyzed modified quantitative-histogram (Q-H) score was used to quantify immunostaining. Main outcome measure(s) The immunoreactions of hepatocyte growth factor (HGF), its receptor, c-Met, vascular endothelial growth factor (VEGF), proliferating cell nuclear antigen (PCNA), and von Willebrand factor (VWF) in eutopic and ectopic endometrium were examined, and their relation with different revised American Society for Reproductive Medicine (r-ASRM) stages and the morphology of endometriosis was evaluated. Result(s) The immunoexpressions of HGF and c-Met were significantly higher in the eutopic endometrium of patients with endometriosis than in that of controls. The Q-H scores of HGF, c-Met, VEGF, PCNA, and microvessel density (MVD) were markedly higher in red peritoneal lesions when compared with other lesions. The Q-H scores did not reveal r-ASRM stage–dependent variation in any of these markers. We observed a significant correlation between the immunoexpressions of HGF, c-Met, and PCNA or microvessel counts. When we combined the Q-H scores of the glandular epithelium and stroma, we found that increased activity of the eutopic endometrium as measured by the immunoreaction of HGF, c-Met, VEGF, PCNA, and MVD was similar to highly active red lesions and was significantly higher than that of controls and other lesions. Conclusion(s) Immunoexpression of HGF and c-Met in the eutopic endometrium of patients with pelvic endometrioisis is possibly useful to predict greater activity of the ectopic endometrium.

Colorectal cancer before the age of 40

Colorectal adenocarcinoma before the age of 40 is uncommon, and its prognosis is controversial, with many studies reporting a worse prognosis than in older patients and others showing no difference. The current study compared two groups of patients who had surgical resection for colorectal adenocarcinoma. The case group was composed of 34 patients younger than 40 (34 +/- 4) years. Detailed pathologic prognosis factors, tumor cell proliferation measured by proliferating cell nuclear antigen, survival, family history, and predisposing conditions were analyzed. Results were compared with a control group constituted of 34 patients older than 65 (75 +/- 6) years matched by gender, cancer site, and Dukes stage. Tumor differentiation, presence of vascular and perineural neoplastic invasion, tumor growth pattern, tumor cell proliferation measured by proliferating cell nuclear antigen count, and survival according to the Kaplan-Meier method were not significantly different between younger and older patients. The only difference between the two groups was a higher prevalence of family history and predisposing conditions for colorectal cancer in younger patients (23 vs. 3 percent; P = 0.03). This case-control study documents that pathologic features and prognosis of colorectal adenocarcinoma are comparable in patients younger than 40 years compared with older patients for identical stages. The higher prevalence of positive family history in younger patients suggests a different genetic background compared with older patients.

The prognostic significance of AgNOR counts and PCNA-positive cell counts in canine malignant lymphomas

The behaviour of canine malignant lymphomas is difficult to predict on the basis of histomorphology alone, but the majority of such tumours are "high-grade" by most classifications. The aim of this study was to examine the prognostic significance of argyrophilic nucleolar organizer region (AgNOR) counts and of proliferating cell nuclear antigen (PCNA)-positive cell counts in canine malignant lymphomas; and to relate the results to the histological grade (Kiel classification) and to the survival time of treated and untreated dogs. Low- and high-grade malignant lymphomas differed significantly in having mean AgNOR counts of 3.3 and 5.7 respectively. Untreated dogs with a mean AgNOR count higher than 5.5 did not survive more than 85 days; the median survival time for this group was 38.5 days. Untreated dogs with a mean AgNOR count lower than 4.0 survived at least 82 days and one dog more than 300 days; the median survival time for this group was 154 days. Treated dogs with a mean AgNOR count higher than 5.5 did not survive more than 126 days; the median survival time for this group was 73.5 days. Treated dogs with a mean AgNOR count lower than 4.0 had a median survival time of 205.5 days and one dog survived 367 days. The mean AgNOR count would seem to be a valuable prognostic marker for canine malignant lymphomas, but PCNA counts had no prognostic significance in relation to the median survival time in treated or untreated dogs.

The evaluation of salivary gland tumours using proliferating cell nuclear antigen.

In order to assess its discriminating and prognostic value, we studied the immunoreactivity for proliferating cell nuclear antigen (PCNA) in tissue from 52 human salivary gland tumours using the murine monoclonal antibody PC10. The PCNA percentage count, namely, the average number of positive nuclei counted per 100 randomly selected tumour cells was recorded for each tumour. Anaplastic carcinoma was used as a positive control and histologically 'normal' salivary gland and tonsil served as a negative control. A PCNA count of 30 per cent was postulated to predict malignancy within a given salivary gland tumour i.e. a PCNA count of 30 per cent or above would indicate malignant potential. This gave a sensitivity of 96.9 per cent and a specificity of 95.2 per cent and a positive predictive value of determining malignancy of 96.8 per cent. We conclude that PCNA immunoreactivity is useful in discriminating between benign and malignant salivary gland tumours and that it may have prognostic value in this diverse group of neoplasms.

Inhibition of progression of chronic puromycin aminonucleoside nephrosis by probucol, an antioxidant.

Oxidants have been shown to be involved in the initiation of chronic puromycin aminonucleoside nephrosis in rats, but it is uncertain whether they have a role in the progression of this disease. Rats given a single internal jugular venous bolus of puromycin aminonucleoside (PA) develop early nephrotic syndrome that subsides after about 28 days followed by a 4-wk period of minimal proteinuria and then the development of focal glomerulosclerosis and increasing proteinuria. Fifty-two rats on a high-cholesterol diet were divided into four groups. Two groups of 16 animals each received a single internal jugular venous bolus of PA. One of these groups was started on the dietary antioxidant, probucol (1% wt/wt), 4 days after the PA injection and continued on it until termination. The remaining rats were given an internal jugular venous injection of an equivolume of normal saline. Five of these animals were also started on dietary probucol 4 days after the saline injection. At 11 days postinjection all animals given PA, whether on probucol or not, developed marked proteinuria with histologically minimal glomerular change and significant increases in intraglomerular monocyte and proliferating cell nuclear antigen counts. Forty-two days after PA injection all PA-injected rats had minimal urinary protein injection and no glomerular changes. At 98 days postinjection rats given PA without probucol developed focal glomerulosclerosis and significant proteinuria compared with PA-injected rats on probucol and those injected with saline (P < 0.05). The probucol-fed PA-injected rats showed no glomerular disease and their urinary protein excretion rates were very similar to those of the saline controls. The results indicate that probucol inhibits the progression of chronic puromycin aminonucleoside nephrosis and are consistent with the suggestion that oxidants are involved in the progression of this entity.

Hepatocyte regeneration in chronic hepatitis C and interferon treatment: Analysis of immunohistological identification of proliferating cell nuclear antigen (PCNA)

To evaluate the usefulness of proliferating cell nuclear antigen (PCNA) immunostaining in the assessment of the efficacy of interferon (IFN) therapy in chronic hepatitis C, we investigated the proliferative activity of hepatocytes in 67 patients with chronic hepatitis C, using this immunostaining method. The percentage of PCNA-positive hepatocytes was 2.4% in patients with chronic persistent hepatitis, 2.5% in those with chronic aggressive hepatitis 2A, and 3.9% in those with chronic aggressive hepatitis 2B. The PCNA count increased with the progression of the liver disease. Patients were classified as complete, partial, and non-responders to IFN; the percentage of PCNA-positive hepatocytes before IFN therapy was 1.6% in the complete responders, 3.9% in the partial responders, and 4.9% in the non-responders. There was a significant negative correlation between the percentage of PCNA-positive hepatocytes and the response to IFN treatment. Thirty-two of 53 cases (60.4%) in which the PCNA labeling index (LI) was less than 5.0 were complete responders compared with 13 of 14 cases (92.9%) in which the PCNA LI was higher than 5.0, representing partial responders or non-responders (P < 0.001). Most complete responders had a low PCNA LI, irrespective of HCV genotype. Our findings indicate that PCNA immunostaining is a simple and reliable index of cell proliferation in liver regeneration, and may be a useful predictor of the response to IFN treatment in chronic hepatitis C.

P53 Protein and Proliferating Cell Nuclear Antigen (PCNA) Expression in Small Round Cell Tumors of Bone and Adjacent Soft Tissue

Sixty small cell tumors of bone and adjacent soft tissue were studied in an attempt to define the incidence of immunohistochemically detectable p53 protein and cor relate these findings with the results of proliferating cell nuclear antigen (PCNA) immunohistochemical staining and mitotic counts. All of the lesions had been for malin-fixed and paraffin-embedded; half were subjected to decalcification prior to processing. The study population included 12 Ewing's sarcomas of bone, 3 atypical Ewing's sarcomas of bone, 3 primitive neuroectodermal tumors of bone, 11 Askin tumors of the thoracopulmonary region, 11 small cell osteosarcomas of bone, 10 mesenchymal chondrosarcomas of bone, and 10 malignant lymphomas involving bone. The patients ranged in age at the time of presentation from 17 to 67 years. Overall, the incidence of p53 positivity was extremely low in these lesions, irre spective of tumor type. Positive nuclear staining with an antibody to p53 was found in none of the 12 Ewing's sarcomas, none of the 3 atypical Ewing's sarcomas, none of the 3 primitive neuroectodermal tumors of bone, 1 of the 11 Askin tumors of the thoracopulmonary region (1.5% of tumor cells positive), 1 of the 11 small cell osteosarcomas (2% of tumor cells positive), 1 of the 10 mesenchymal chondrosar comas of bone (7% of tumor cells positive), and 2 of the 10 malignant lymphomas involving bone (0.5% and 1% of tumor cells positive, respectively). The majority of tumors showed PCNA positivity within the tumor cells, although the incidence of PCNA positivity within the histologic types varied greatly; in general, the higher PCNA counts corresponded to higher mitotic counts within the individual lesions. The present study did not demonstrate any correlation between mutant p53 accu mulation detected by immunohistochemistry and tumor type, and so it is unlikely that p53 positivity will prove to be of great use in the differential diagnosis of these lesions. A correlation between p53 positivity and PCNA staining or mitotic activity was not apparent. Int J Surg Pathol 2(4):259-268, 1995

Assessment of proliferating-cell nuclear antigen immunostaining in soft-tissue tumours: relationship to histological grade and mitotic activity.

The proliferative activity in 70 cases of soft-tissue tumours was estimated immunohistochemically using the monoclonal antibody PC-10, which recognizes proliferating-cell nuclear antigen (PCNA) in paraffin sections. The PCNA index (i.e. the percentage of positive neoplastic nuclei) and to a lesser degree the PCNA count (i.e. the number of positive neoplastic nuclei per ten high-power fields) positively correlated with the malignancy grade (PCNA index: P < 0.001; PCNA count: P < 0.01). However, the range of values of PCNA index and PCNA count was similar between benign and grade I tumours. A statistically significant positive correlation was also established between PCNA index and PCNA count on the one hand and mitotic count on the other, but the correlation coefficient was low (r = 0.351, P < 0.01, and r = 0.290, P < 0.05 respectively). These results indicate that PCNA immunostaining may successfully be used as an adjunct to the conventional histopathological parameters in assessing the malignancy grade in soft-tissue tumours although it is of limited value in distinguishing between benign and grade I tumours.

Proliferating cell nuclear antigen counts as markers of cell proliferation in head and neck cancer.

Proliferating cell nuclear antigen (PCNA) expression was investigated immunhistochemically in 53 squamous cell carcinomas of the head and neck. PCNA is a 36-kDa nuclear protein associated with the cell cycle. Results were compared with Ki67 counts, with this latter marker used to demonstrate proliferative compartments. Overall, the PCNA and Ki67 labelling index showed a similar distribution pattern in normal and tumor tissue. A strong correlation was found to histological differentiation. There was no significant difference between PCNA and Ki67 counts (r = 0.8), and PCNA immunostaining allowed assessment of proliferative activity in the head and neck cancers studied. The application of formalin-fixed, paraffin-embedded tumor material demonstrated the advantage of this method and showed that it is an excellent alternative to Ki67 counts.

Small gliomas: clinicopathological study.

The clinicopathological characteristics of small gliomas were investigated in nine patients with gliomas less than 2 cm in diameter. The tumor histology and proliferative activity were also examined. Three tumors occurred in the white matter and six at the corticomedullary junction. Histological diagnosis, Daumas-Duport's grading, bromodeoxyuridine labeling index, and proliferating cell nuclear antigen counts were well correlated. Most small gliomas were histologically benign, localized, and removable if not in eloquent areas of the brain. However, some cases showed high-grade glioma and invasive character at the early stage, resulting in a poor outcome.

Assessment of rejection in orthotopic human heart transplantation using proliferating cell nuclear antigen (PCNA) as an index of cell proliferation.

Myocardial biopsies taken during the management of cardiac transplantation were stained for proliferating cell nuclear antigen (PCNA). Counts of PCNA-positive interstitial cells were compared, in retrospect, with the reported histological grade of rejection. Biopsies without rejection had negligible numbers of PCNA-positive cells. Ascending grades of rejection were paralleled by an increase in the number of PCNA-positive cells [grade 1, 13 +/- 35 (mean +/- SD); grade 2a, 38 +/- 40; grade 2b, 91 +/- 75; grade 3, 170 +/- 78]. While highly significant, in statistical terms, the overlap in the counts between different grades means that prediction of rejection from the PCNA count alone is not feasible. Biopsies graded as 0 or 1 and which immediately preceded more severe rejection episodes showed no increase in PCNA-positive cells. The majority of PCNA-positive cells are fibroblasts, although in grade 2b and 3 rejection a small population of PCNA-positive T lymphocytes occurs. PCNA staining is also seen in cardiac myocytes immediately after transplantation, during rejection episodes, and late after transplantation in the absence of rejection. The positive PCNA staining of cardiac myocytes probably reflects DNA synthesis that occurs with the shift toward polyploidy in hypertrophy.