Argyrophilic Nucleolar Organizer Regions Score Research Articles

Association of Proliferative Indices With Various Grades of Breast Carcinoma.

Various grades of breast carcinoma and proliferative indices used as nuclear protein Ki-67 and argyrophilic nucleolar organizer regions (AgNOR) are being compared to each other. In this observational cross-sectional investigation, 42 breast biopsies from questionable breast areas were collected and preserved in formalin and paraffin before the tissue blocks were made. A thorough medical history regarding the breast tumor and thorough physical examination results were recorded. Two sections were produced, one stained with an immunohistochemical marker called Ki-67 and the other with a unique stain called AgNOR. Grade I in Nottingham was found to be highest in subjects with Ki-67 1%, grade II in subjects with Ki-67 1-10%, and grade III in subjects with Ki-67>10%. Therefore, a higher Ki-67 score and a higher Nottingham grade were more closely associated. The mean AgNOR score was determined to be highest in Nottingham grade III and lowest in Nottingham grade I. In contrast to grade I and grade II of carcinoma (CA) breast, where there was no statistically significant association between Ki-67 and AgNOR, grade III of CA breast showed a statistically significant link between Ki-67 and AgNOR. Proliferation has been identified as a distinctive feature of cancer and as a key factor in the prognosis of the disease.

The Effect of Thelison Use in The Etiology of Lung Disorders among Homeless People

Background: Thelison, the adhesive synthetic material that bind surface together, is widely used in industry and domestic purpose along with epoxy, glue and putty. The aim of this study was to detect the effect of thelison use in the etiology of lung disorders among homeless people in Khartoum State, Sudan.Materials and method: This was a descriptive cross sectional study conducted in homeless people in Khartoum State.Sputum smears samples from 80 alcohol fixed homeless thelison user were collected. After the collection of the sample, Argyrophilic Nucleolar Organizer Region (AgNOR) technique and papanicolaou (PAP) stains were applied to each sample. A questionnaire to obtain essential data about respondents was also provided for each participant.Results: Participant’s ages range was 10-37, while the mean age was 17. The range of participants duration of use per years was 1-10, while the mean was 3 years. Number of thelesion dose use per day was ranged between 1-10, while the mean was 5 years. The majority of participants (80%) showed no cellular change and 20 % showed chronic inflammation. Results showed that 31 of the study population were males (77.5%), while the female population of the study was 9 (22.5%). The mean of AgNoR score was ranged between 1-4, while the mean AgNOR score was 1. AgNoR showed insignificant association with gender, duration and number of thelesion dose used per day (p>0.05), but showed significant association with cytomorhpological and age (p<0.05).Conclusion: AgNoR score showed insignificant association with gender, duration and number of thelesion dose used per day (p>0.05), but showed significant association with cytomorhpological and age (p<0.05).Keywords: AgNoRs score, cytological changes, sputum, homeless thelison users

Prognostic Validity of AgNOR in Pleomorphic Adenoma

Since the argyrophilic nucleolar organizer region (AgNOR) technique has successfully distinguished various grades of malignancies and enabled prognostic assessment, this paper traces its prognostic validity in predicting the behavior of pleomorphic adenoma (PA). Ten cases of recurrent PAs were compared, on the one hand, where AgNOR score of area fraction was measured before and after recurrence. The same findings were contrasted to ten cases of normal glandular mucosa. Diagnosing both pleomorphic adenomas was based on clinical and histological records of the archival submitted cases. The data were statistically analyzed by t-test, ANOVA, Tukey and Pearson's tests. The study concludes that AgNOR score lack the prognostic validity on testing PA, in terms of distinguishing and or predicting recurrence.

Patterns of nucleolar organizer region in urinary bladder cancer in view of 2004 WHO classification of urothelial neoplasms

Context: The last decade has seen a tremendous upheaval since introduction of WHO-1998 classification (modified in 2004) of bladder carcinoma, which only recently seems to have settled down. We compare both old (WHO-1973) and new (2004) classifications with argyrophilic nucleolar organizer region (AgNOR) score. Aims: To compare WHO/ISUP (World Health Organization/International Society of Urologic Pathologists) consensus classification-2004 and WHO-1973 classification of bladder carcinoma and to establish correlation between AgNOR scores and various grades of carcinoma bladder. Materials and Methods: Sixty H and E (Hematoxylin and Eosin) stained sections of transitional cell carcinoma bladder were graded according to WHO/ISUP (2004) and WHO (1973) classification systems and the results were compared. This was then followed by AgNOR staining of the paraffin-embedded sections and subsequent AgNOR scores were calculated and given as total score, extranucleolar score, intranucleolar score, and nucleolar score. These were correlated with the grade of the tumor. Statistical Analysis Used: ANOVA test. Results: It was observed that the two grading systems, WHO/ISUP (2004) and WHO (1973) correlated with each other. Also it was observed that the total AgNOR score and extranucleolar AgNOR score increased with the grade of the tumor and thus increasing scores carry significance in assessment of the grade of the tumor. However, intranucleolar and nucleolar scores showed variable degree of overlap in various grades of the bladder tumors. Conclusions: WHO/ISUP (2004) classification correlated well with the WHO (1973) classification. The total AgNOR score was found to be helpful in assessment of the grade of the tumor but the distribution of the AgNOR carried little significance.

Canine Subcutaneous Mast Cell Tumour: Diagnosis and Prognosis

The aim of this study was to characterize the pathology and clinical outcome of the subcutaneous variant of canine mast cell tumour. Fifty-three cases satisfying the inclusion criteria were selected from the pathology archive of the College of Veterinary Medicine, University of Tennessee. Referring veterinarians provided information on outcome. These dogs had a median age of 9 years (range 3-17 years). After characterizing tumours histologically, nuclear expression of proliferating cell nuclear antigen (PCNA) and Ki67 (MIB-1 clone) was determined immunohistochemically and mast cell origin was confirmed with c-Kit staining. Counts of argyrophilic nucleolar organizer regions (AgNOR) were determined by silver staining. Nuclear labelling was counted in 100 tumour cells. Margins were recorded as incomplete in 66% of dogs, and metastases occurred in 6% of dogs. The estimated minimum mean survival time from date of diagnosis was 1199 days, ranging from 55 to >1780 days. The median scores from immunohistochemical labelling were PCNA 0.05 and Ki67 0.03 per 100 tumour cells. The median score for AgNOR staining was 1.25 per 100 tumour cells. The patterns of c-Kit expression included membranous labelling in 20 tumours, stippled cytoplasmic labelling in 23 tumours and diffuse cytoplasmic labelling in 10 tumours. Age (r=-0.61, P=0.14) and AgNOR score (r=-0.58, P=0.17) had moderate, but non-significant, negative associations with survival. PCNA (r=-0.32, P=0.47), Ki67 (r=-0.22, P=0.64) and c-Kit immunolabelling was not associated with survival. The subcutaneous variant of canine mast cell tumour is distinct in having features of intermediate histological grade and extended mean survival times, suggesting a slightly better long-term prognosis than for higher grade dermal variants. Expression of nuclear proliferation markers is not associated with outcome.

Diagnostic significance of FDG-PET and argyrophilic nucleolar organizer regions (AgNORs) in oral squamous cell carcinoma

Neoadjuvant chemoradiotherapy has played an important role in preserving organs and functions in patients with oral squamous cell carcinoma (SCC). To determine whether further operation after chemoradiotherapy is necessary, accurate evaluation of residual tumour cells after treatment is essential. We investigated the usefulness of regional fluorodeoxyglucose (FDG) uptake in combination with histochemical expression of AgNORs obtained from pretreatment biopsy specimens to assess residual tumours after intraarterial chemoradiotherapy. Standardized uptake value (SUV: pretreatment, pre-SUV; post-treatment, post-SUV) was calculated to determine the changes of FDG uptake in the tumour before and after chemoradiotherapy. Four out of 20 patients with oral SCCs showed residual tumour cells after treatment. Tumours with higher post-SUVs (⩾4.0) and higher AgNORs scores (⩾5.0) had significantly higher incidences of residual viable tumour cells after chemoradiotherapy ( P=0.001). The results suggest that the combination of FDG-PET with AgNORs score is an excellent index for determining the optimal management of each patient following neoadjuvant chemoradiotherapy.

Evaluation of argyrophilic nucleolar organizer regions (AgNORs) in differentiated thyroid carcinoma as an indicator for disease recurrence.

Differentiated thyroid carcinoma (DTC) is one of the least aggressive cancers of the human malignancies, however, recurrent disease is occasionally found and is difficult to determine the risk for recurrence only by clinicopathological features. In the present study, we investigated argyrophilic nucleolar organizer regions (AgNORs) score, a well-known indicator for proliferative potential of the cancer cells, in 89 cases of DTC. In tumors with capsular invasion, or with extended contra-lateral lymph node metastasis, AgNORs score was significantly higher than in tumors without them, while was not correlated with age or gender of the patients, nor the histological type of the tumor (papillary or follicular). Disease recurrence was observed in one-third of the patients in high AgNOR score group (scored more than mean value +/- standard deviation) when the patients were divided into four groups according to the AgNOR score, and a significantly higher risk for disease recurrence was demonstrated in those cases with high AgNOR score than cases with lower score. These results clearly indicated the usefulness of AgNOR score in selecting the patients at high risk for disease recurrence. All 5 recurrent cases displayed local recurrence in high AgNOR score group, while 5 of 7 cases with low AgNOR score did in the manner of hematological metastasis. In conclusion, this method was technically simple and accurate giving an exact value in individual cases. Thus, we believe AgNOR score might be clinically applicable as a useful indicator for disease recurrence in DTC.

ARGYROPHILIC NUCLEOLAR ORGANIZER REGION IN PROLIFERATING CELL HAS A PREDICTIVE VALUE FOR LOCAL RECURRENCE IN SUPERFICIAL BLADDER TUMOR

Purpose It has been shown in many carcinomas that the proliferation rate and number of argyrophilic nucleolar organizer regions (AgNOR) are associated with tumor aggressiveness. However, in bladder tumor the significance of the correlation between the number of AgNOR and tumor behavior remains controversial. Therefore, it would be helpful if a new technique could be developed that would allow for more accurate AgNOR counting in association with tumor behavior. We established the simultaneous staining technique of AgNOR with Ki-67 labeling to reveal the significance of AgNOR count in superficial bladder tumor. Materials and Methods A total of 50 paraffin sections of superficial bladder tumor were stained with AgNOR and Ki-67 (MIB-1). The numbers of AgNORs in proliferating (MIB-1 positive) or resting (MIB-1 negative) cells were counted from a total of 100 nuclei. Correlations between MIB-1 associated AgNOR count and clinicopathological parameters were statistically analyzed. Results The AgNOR count in proliferating cells was significantly higher than that in resting cells (p <0.01), and the count significantly increased with tumor grade (p <0.01). Based on recurrence-free survival analyses the local recurrence rate was significantly higher in patients with high proliferating cell NOR but not for those with resting or whole cells. However, no AgNOR score helped to select patients at high risk for disease progression. Conclusions Proliferating cell NOR had a predictive value for local recurrence in patients with superficial bladder tumor.

Expression of nm23-H1 in colorectal cancer: no association with metastases, histological stage, or survival.

The correlations of nm23-H1 expression in primary cancer lesions with the already confirmed 14 prognostic variables and survival were examined in 52 advanced colorectal cancer patients, because the clinical roles of nm23-H1 expression in the cancer lesions remain controversial. An immunohistochemical expression of nm23-H1 was found in 23 lesions (positive group) but not found in 29 lesions (negative group). No significant difference between the positive and negative groups was found according to 12 clinicopathological variables including vascular invasion, lymph node and liver metastases, and histological stage. The carcinoembryonic antigen levels (21.5+/-33.4 ng/ml) of the draining venous blood and argyrophilic nucleolar organizer regions score (3.35+/-1.36 per nucleus) of the cancer cells in the positive group were not significantly diffeent from those (34.1+/-102.9 ng/ml and 3.32+/-1.00 per nucleus, respectively) in the negative group. In addition, no significant difference was found in the survival curves or the 5-year survival rates of the positive and negative groups. From these results, it may be concluded that the nm23-H1 expression was not associated with the aforementioned prognostic variables and the prognosis of advanced colorectal cancer patients.

Evaluation of argyrophilic nucleolar organizer region and proliferating cell nuclear antigen in colorectal cancer.

Information on cellular proliferation is gaining importance for predicting prognosis in several cancers. To clarify the clinicopathological significance of argyrophilic nucleolar organizer region (AgNOR), proliferating cell nuclear antigen (PCNA), and DNA ploidy pattern, we studied their correlations with clinicopathological factors in colorectal cancer. Fifty-two patients with colorectal cancer were examined by AgNOR staining, immunohistochemical study of PCNA expression, and DNA flow cytometry. The AgNOR score and the PCNA labeling rate (PCNA LR) were significantly higher in patients with deep invasion (P = 0.0072, P = 0.0355), liver metastasis (P = 0.0022, P = 0.0001), and Dukes D classification (P = 0.0002, P = 0.0001) than in patients without these factors. In patients with high AgNOR score (>3.83) or with high PCNA LR (>48.8), prognosis was significantly worse (P = 0.0002, P = 0.0123) than in those with low AgNOR score (<3.83) or in those with low PCNA LR (<48.8), respectively. No significant association was observed between AgNOR score and PCNA LR. Combined analysis revealed that the survival curve for patients with high AgNOR score and high PCNA LR was significantly lower (P = 0.0156) than that for patients with high AgNOR score and low PCNA LR. There was no significant correlation between DNA ploidy pattern and clinicopathological findings. PCNA LR and AgNOR score were correlated not only with local progression but also with metastasis. Their determination provided useful prognostic information, and these parameters are probably independent. Their simultaneous determination was useful for accurate evaluation of prognosis. The value of DNA ploidy pattern was uncertain.

Usefulness of Argyrophilic Nucleolar Organizer Regions Score to Differentiate Suspicious Malignancy in Pulmonary Cytology

Pulmonary cytologic specimens reported as "suspicious for malignancy" pose problems in clinical management. Silver staining for argyrophilic nucleolar organizer regions (AgNOR) has proved useful in making a cytopathologically differential diagnosis between benign and malignant cells. This study aimed to evaluate the usefulness of AgNOR score in the diagnosis of pulmonary cytologic specimens deemed inconclusive by conventional staining methods. Pulmonary cytologic specimens initially reported as suspicious for malignancy with Papanicolaou or May-Grünwald-Giemsa (MGG) staining obtained from 35 proved cases were destained then restained using the AgNOR technique. Another 35 cases with clear cytologic diagnosis were also examined for comparison. The median number of dots, defined as the AgNOR score, was used to differentiate malignant from benign specimens. Malignant cases had significantly higher AgNOR scores than benign ones (p<0.001). There were no significant differences among smears previously stained with Papanicolaou or MGG method, among specimens obtained via bronchoscopic brushing, fine-needle aspiration of lung or pleural effusion, or among subgroups of malignant diseases. Based on the results of our previous study, the cutoff value of the AgNOR score to differentiate benignancy from malignancy was set at 6. At this setting, the sensitivity and specificity of AgNOR score were 88% and 80%, respectively, in aiding a differential diagnosis of pulmonary cytologic specimens initially classified as suspicious for malignancy. For those cases with a clear cytologic diagnosis, the sensitivity and specificity of AgNOR score were 92% and 100%, respectively. For all cases, the sensitivity of AgNOR score was 90% and the specificity was also 90%. The AgNOR score is of value in aiding a differential diagnosis between benign and malignant lesions in pulmonary specimens with equivocal cytologic features.

The argyrophilic nucleolar organizer region (Ag-NOR) score in invasive ductal carcinoma of the pancreas.

The biological malignancy of pancreatic carcinoma was evaluated by determining the score of the argyrophilic nucleolar organizer region (Ag-NOR) in resected specimens, and comparing it with the various clinicopathological factors and long-term results of 38 patients who underwent surgical resection for invasive ductal carcinoma of the pancreas between 1977 and 1992, in whom the Ag-NOR could be stained. The Ag-NOR analysis of pancreatic carcinomas from the 38 patients resulted in a mean Ag-NOR score of 3.82 +/- 0.62, which was significantly (P < 0.01) higher than the mean score of 1.72 +/- 0.28 observed in normal pancreatic tissues obtained from 20 of these patients. The mean Ag-NOR score significantly increased in patients with anterior capsular infiltration (s) (P < 0.01), posterior tissue infiltration (rp) (P < 0.01), and lymph node metastasis (n) (P < 0.05) compared to those without these factors. The rate of curability A or B was only 13.0% in patients with a high Ag-NOR score of > or = 3.80 in comparison to 66.7% in those with a low Ag-NOR score of < 3.80. The 3-year survival rate was significantly (P < 0.05) higher in the low Ag-NOR score group than in the high Ag-NOR score group. These results suggest that the Ag-NOR score can serve as an indicator of the biological malignancy of pancreatic carcinoma, and of the patients' prognosis.

Argyrophilic Nucleolar Organizer Regions of Breast Tumors: Assessment of Aspirated Cytological Materials.

To investigate the correlation between argyrophilic nucleolar organizer regions (Ag-NORs)and the malignant potential of breast tumors, we analyzed Ag-NORs of cytological specimens obtained from 190 patients with various types of breast disease by preoperative aspiration biopsy. The average number of Ag-NORs per nucleus was defined as the Ag-NOR score. The Ag-NOR score was 5.7 +/- 1.7 in the group of women with breast carcinoma (n =70), 2.6= 0.4 in the group with fibroadenoma (n= 54) and 2.9= 0.6 in the group with mastopathy (n= 66). The level was significantly higher in breast carcinoma than in each benign disease(P < 0.001 in both cases). The score was 6.5 +/- 2.3 in the group with four or more metastatic lymph nodes (n = 21), 5.2+/- 1.2 in the group with one to three metastatic lymph node (n= 10) and 5.1+/- 1.1 in that with no metastatic lymph node (n = 39);the score was significantly (P < 0.05) higher in the groups with four or more metastatic lymph modes than in the other groups, respectively. Thus, a correlaton was observed between the Ag-NOR score and lymph node status. These data suggest that a higher Ag-NOR score reflects high-grade malignancy.

Nucleolar Organizer Regions: A New Prognostic Factor for Upper Tract Urothelial Cancer

Purpose The prognostic significance of argyrophilic staining in the nucleolar organizer regions was studied in 63 patients with primary urothelial tumors of the renal pelvis and ureter. Materials and Methods Silver staining of paraffin embedded sections was performed using a 1-step technique. Results The prognosis for patients with a mean number of argyrophilic nucleolar organizer region proteins per nucleus (argyrophilic nucleolar organizer region score) of 8 or greater was significantly worse than that for patients with a score of less than 8 (p less than 0.001). Conclusions Argyrophilic nucleolar organizer region score is a new prognostic factor in primary urothelial tumors of the renal pelvis and ureter, and it is particularly useful for patients with invasive tumors.

Number of argyrophilic nucleolar organizer regions is a good indicator of lymph node metastasis in patients with esophageal carcinoma.

The number of argyrophilic nucleolar organizer regions (AgNORs) was evaluated as a predictor of lymph node metastasis in 45 patients who had undergone resection of advanced squamous cell carcinoma of the esophagus. The mean AgNOR score of carcinomas was 5.0 +/- 1.8, and it was greater than that of normal esophageal epithelium adjacent to a carcinoma (2.3 +/- 0.5, P < 0.001). The AgNOR score of tumors from 26 patients with lymph node metastasis was 6.1 +/- 1.6, and it was greater than that of tumors from 19 patients without lymph node metastasis (3.7 +/- 1.0, P < 0.001). The AgNOR scores of metastatic lymph nodes (4.9 +/- 1.5) from 26 patients with lymph node metastasis were closely related to the number of metastatic lymph nodes of individual patients (r = 0.582, P < 0.0001). The 3-year survival rate in patients with low AgNOR scores (AgNOR score < 5, n = 22) was 56.2%. By contrast, that in patients with high AgNOR scores (AgNOR score > or = 5, n = 20) was only 13.1%. There was a statistically significant difference between the two survival curves (P < 0.05). These results indicate that the AgNOR score is a good indicator of lymph node metastasis and suggest that it might also be a useful prognostic marker in patients with esophageal cancer.

Argyrophilic nucleolar organizer regions (AgNORs) in breast lesions--a study on fine needle aspirates.

Nucleolar organizer region-associated proteins are argyrophilic and may be visualized by silver stains. This study is a prospective analysis of the AgNOR counts in a range of breast lesions seen in fine needle aspiration (FNA) smears. A total of 72 cases including ductal carcinomas (n = 24), fibrocystic change (n = 24), fibroadenoma (n = 16), proliferative lesions (n = 4) and miscellaneous lesions (n = 4) were studied. The ductal carcinomas showed higher AgNOR counts (mean 16.63 +/- 7.09) compared with the non-malignant lesions (mean 6.39 +/- 1.96), the difference being statistically significant. A cut off AgNOR score of 11 could reliably discriminate malignant from non-malignant lesions. In ductal carcinomas, the AgNOR scores showed a tendency to increase with higher grades of malignancy. There was no correlation with the presence or absence of lymph node metastases at the time of presentation. It is concluded that AgNOR counting, although tedious, is inexpensive, and provides useful information regarding cellular proliferation, and can supplement information obtained by more sophisticated techniques.

The significance of nucleolar organizer region (AgNOR) score in predicting meningioma recurrence

Argyrophilic nucleolar organizer region (AgNOR) has been demonstrated in recent studies of human brain tumors, including meningiomas. The authors used this technique in meningiomas to analyze whether the mean numbers of AgNOR per nucleus (AgNOR score) are related to the recurrence rate or the proliferative potential of meningiomas. AgNOR staining technique was applied to paraffin-embedded sections of 60 meningiomas. Twenty of these specimens also were investigated immunohistochemically with monoclonal antibody (MoAb) against DNA polymerase alpha (Pol.alpha) and with MoAb Ki-67 to compare the AgNOR score with the proliferative potential. There was a statistically significant difference between the AgNOR score in nonrecurrent meningiomas (2.48 +/- 0.73) and recurrent histologically benign meningiomas after gross total resection (3.20 +/- 0.96) (P < 0.02). The recurrence rate of tumors after gross total resection with an AgNOR score of 3.0 or more was significantly higher than that with an AgNOR score of less than 3.0. The AgNOR score did not always correlate with Pol.alpha or Ki-67 score; the AgNOR score of malignant meningioma that had high proliferative score was not always high. This study indicates that AgNOR staining appears to be a simple and useful method for estimating the probability of histologically benign meningioma recurrence even after gross total resection.

PROGNOSIS FOR GASTRIC-CANCER INVADING THE SEROSA EVALUATED BY ARGYROPHILIC NUCLEOLAR ORGANIZER REGION STAINING

Eighty-five patients with gastric cancer invading the serosa following curative resection were analyzed with respect to clinicopathological features and prognosis by staining the argyrophilic nucleolar organizer region (AgNOR). The AgNOR was counted in 200 cancer cells per tissue and the mean number per cell was identified as the AgNOR score. The AgNOR count varied from 1.79 to 5.71, while the mean value was 2.92 +/- 0.74 and the median score was 2.95. The patients were divided into two groups: the AgNOR high group (AgNOR count greater-than-or-equal-to 2.95, n=44) and the AgNOR low group (AgNOR count < 2.95, n=41). There were no differences between the two groups with respect to sex, age, tumor size, location of the tumor, macroscopic appearance, histological differentiation, growth pattern and vascular involvement. The lymphatic advancement of tumor cells was prominent and the rate of lymph node metastasis was higher (p<0.01) in patients in the AgNOR high group. The AgNOR high group had a higher rate of recurrence and also had a variety of recurrence styles. The prognosis of the AgNOR high group was poorer, with statistical significance (p<0.01), while the 10-year survival rate was 35.3% for the AgNOR high group and 77.2% for the AgNOR low group. Therefore, the AgNOR count was found to be closely related to tumor advancement and thus is considered to have a predictive value for the prognosis of patients with gastric cancer invading the serosa after curative resection.

A correlation of argyrophilic nucleolar organizer regions with stages of hepatocellular carcinoma.

Recently, the number and morphologic features of argyrophilic nucleolar organizer regions (AgNOR), silver-stained nucleolar organizer regions, were thought to reflect the cellular proliferative activity and the grade of malignancy. In liver diseases, it has been reported that AgNOR scores for hepatocellular carcinomas (HCC) were significantly higher than those for benign and borderline lesions; the scores increased with histologic tumor grade. Using the colloid silver staining technique to reveal AgNOR, 64 liver biopsy specimens with HCC were examined of which 14 had Stage I disease, 20 had Stage II, 14 had Stage III, 10 had Stage IVA, and 6 had Stage IVB (by International Union Against Cancer criteria). AgNOR in nuclei were divided into two types. Type 1 (T1-NOR) contained large and medium-sized brown dots with well-defined margins. Nucleoli were included in this type. Type 2 (T2-NOR) had fine black single dots and clusters without well-defined margins outside T1-NOR, but within the nucleus. The size and irregularity of T1-NOR changed with the progress of cancerous stages. The increase in the number of T2-NOR contributed preponderantly to the overall increase in Ag-NOR scores. These results suggest that HCC with smaller and/or irregular T1-NOR in combination with high T2-NOR scores have a more aggressive potential. AgNOR may be useful indicators for evaluating the progress of HCC.