Roberto Navone

The impact of liquid-based oral cytology on the diagnosis of oral squamous dysplasia and carcinoma

Objective:  Even though diagnostic oral exfoliative cytology is a useful, economical and practical tool in the diagnosis of oral dysplasia and carcinoma, it is not yet extensively used. The results of conventional exfoliative and liquid‐based diagnostic cytology in oral potentially malignant lesions (PML) are herein reported and compared with the histological diagnosis.

Biologic differences between noninvasive papillary urothelial neoplasms of low malignant potential and low-grade (grade 1) papillary carcinomas of the bladder.

We investigated the expression of oncogenes p53, c-erbB-2, and bcl-2 and cell proliferative activity in 62 newly diagnosed superficial pTa papillary bladder tumors. Based on the 1998 World Health Organization/International Society of Urological Pathology (WHO/ISUP) and 1999 WHO classifications, 19 were urothelial neoplasias of low malignant potential (LMP) and 43 low-grade (grade 1) papillary carcinomas. All the patients underwent transurethral resection and were followed up to 97 months; 42 had recurrences. Initial biopsies were tested for p53, c-erbB-2, and bcl-2 proteins using DO7, CB11, and bcl-2 124 monoclonal antibodies. Cell proliferation was assessed by MIB-1 mAb and mitotic count. LMP had significantly lower MIB-1 (p = 0.002) and p53 immunopositivity (p = 0.03), mitotic count (p = 0.006), and recurrence rates (p = 0.04) than did grade 1 cases, whereas no difference was observed for c-erbB-2 and bcl-2 expression. The median disease-free survival for LMP was 76 months but only 15 months for grade 1 cases (p = 0.002). Although the cohort is small, the results indicate that the distinction between LMP and low-grade (grade 1) papillary urothelial neoplasias, as proposed by the 1998 WHO/ISUP and 1999 WHO classifications, reflects different biologic activity and clinical behavior; however, a long-term follow-up is advisable also for patients with LMP.

Lymphoid Nodules and Nodular Lymphoid Hyperplasia in Bone Marrow Biopsies

Out of 2,474 bone marrow biopsies we have observed 330 cases (13.3%) with presence of lymphoid nodules (LN). LN were frequent in old age (24.6% over 80 years), in females (17%) and in some diseases, such as rheumatoid arthritis and systemic lupus erythematosus (73.7% of the cases), partial aplasia (34%), hypersplenism (30.4%), hemopoietic dysplasia (25%), chronic renal failure (20.4%), polycythemia vera (20.2%), idiopathic thrombocytopenic purpura (18.8%), acute leukemia (17.7%). Nodular lymphoid hyperplasia of the bone marrow was found especially in systemic autoimmune diseases (26.3%), hypersplenism (9.8%), preleukemia (7.3%) and acute leukemia (4.2%). The presence of excessive medullary LN could indicate a bone marrow microenvironment damage, possibly of autoimmune origin.

DNA aneuploidy and dysplasia in oral potentially malignant disorders: Association with cigarette smoking and site

To date there are still no reliable biomarkers for oral potentially malignant disorders (PMDs) to predict the risk of progression to squamous cell carcinoma (SCC). Within a prospective clinical trial of patients with PMDs, DNA content flow cytometry (DNA FCM) was evaluated for 60 PMDs using fresh samples obtained by a dermatological curette. There were 6/42 PMDs without dysplasia, but with DNA aneuploidy, versus 8/18 with both dysplasia and aneuploidy (p=0.02). When the tongue and the buccal mucosa, the two most common sites in the present series of cases were compared, dysplastic PMDs were mainly located on the tongue (p=0.01). Tobacco smokers, who preferentially developed PMDs in the buccal mucosa at a younger age than non-smokers (p=0.002), had fewer dysplastic PMDs than did non-smokers (p=0.01). Dysplasia was significantly linked to DNA aneuploidy (p=0.03) in smokers. The present data suggest that aneuploidy is an early event in oral carcinogenesis and that the influence of tobacco varies according to subsite and patient age. When DNA FCM of PMD samples are obtained by curette scraping, extensive areas can be covered with a minimally invasive, rapid, inexpensive procedure. Moreover DNA FCM of these samples appears easy amenable to routine analysis. Further research on larger numbers of PMDs should be carried out to determine whether DNA FCM plays a role in the prediction of risk of PMD transformation.

Argyrophilic nucleolar organizer region counts and proliferating cell nuclear antigen scores are two reliable indicators of survival in pharyngeal carcinoma

SummaryThe proliferative activity of pharyngeal carcinoma has been investigated by means of monoclonal antibody PC10 against proliferating cell nuclear antigen (PCNA/cyclin) and argyrophilic nucleolar organizer region (AgNOR) analysis in formalin-fixed, paraffin-embedded biopsies from 45 primary squamous and undifferentiated carcinomas, prior to therapy.The correlation between AgNOR counts and PCNA(PC10) scores was highly significant (r=0.73;P<0.0001) as determined by Pearsons correlation coefficient. Moreover, the univariate Kaplan-Meier survival analysis showed a significant correlation between 3- and 5-year survival rates and the mean AgNOR number per tumour cell (P=0.0003) or the percentage of PCNA(PC10)-positive cells (P=0.0001).Our results indicate that both AgNOR counts and PCNA(PC10) scores are reliable markers of the proliferative activity of pharyngeal carcinoma in small, routinely processed biopsies, in which they can allow simultaneous evaluation of the histology and tumour cell kinetics.

Cell proliferation indices, morphometry and DNA flow cytometry provide objective criteria for distinguishing low and high grade bladder carcinomas

Argyrophilic nucleolar organizer region (Ag-NOR) analysis, proliferating cell nuclear antigen (PC-NA/PC10) and MIB-1 immunohistochemistry, nuclear morphometry and DNA flow cytometry have been performed on formalin-fixed, paraffin-embedded biopsies from 50 patients with transitional cell carcinoma of the urinary bladder. The mean AgNOR count was 6.01 for the 17 grade 1 (G1), 7.59 for the 21 G2 and 13.33 for the 12 G3 carcinomas (p<0.001). The mean PCNA score was 15.03% for G1, 24.04% for G2 and 40.01% for G3 cases (p<0.001). The mean MIB-1 score was 11.31% for G1, 17.09% for G2 and 34.47% for G3 carcinomas (p<0.001). The mean nuclear area was 35.53 μm2 for G1, 38.65 μm2 for G2 and 83.62 μm2 for G3 cases (p<0.001). Aneuploidy rates were significantly higher (91.7%) in G3 than in G2 (42.9%, p<0.01) or G1 cases (47.1%, p<0.05) but not different for G1 versus G2 cases (p=0.94). While many overlaps of values were seen between G1 and G2 tumours, no overlaps were found between G3 and G1/G2 tumours. Significant differences of values were also found between pTa and invasive tumours (p<0.0001 for AgNOR count and PCNA score; p<0.001 for MIB-1 score and mean nuclear area; p<0.01 for DNA ploidy); however many overlaps were seen. Our findings indicate that the quantitative parameters obtained with different methods are associated with histological grade of bladder urotheliomas and may improve the grading reproducibility. In addition, the absence of overlaps between G3 and G2/G1 carcinomas supports the tendency to classify bladder urotheliomas in only two categories of malignancy.

Oral cancer genesis and progression: DNA near-diploid aneuploidization and endoreduplication by high resolution flow cytometry

Oral potentially malignant lesions (OPMLs) with dysplasia and aneuploidy are thought to have a high risk of progression into oral squamous cell carcinomas (OSCCs). Non-dysplastic “oral distant fields” (ODFs), characterized by clinically normal appearing mucosa sited at a distance from co-existing OPMLs, and non-dysplastic OPMLs may also represent an early pre-cancerous state. ODFs, OPMLs without and with dysplasia and OSCCs were investigated by high resolution DNA content flow cytometry (FCM). ODFs and OPMLs without dysplasia were DNA aneuploid respectively in 7/82 (8.5%) and 25/109 (23%) cases. “True normal oral mucosa” and human lymphocytes from healthy donors were DNA diploid in all cases and were used as sex specific DNA diploid controls. Dysplastic OPMLs and OSCCs were DNA aneuploid in 12/26 (46%) and 12/13 (92%) cases. The DNA aneuploid sublines were characterized by the DNA Index (DI ≠ 1). Aneuploid sublines in ODFs and in non-dysplastic and dysplastic OPMLs were near-diploid (DI < 1.4) respectively in all, 2/3 and 1/3 of the cases. DNA aneuploid OSCCs, instead, were characterized prevalently by multiple aneuploid sublines (67%), which were commonly (57%) high-aneuploid (DI ≥ 1.4). DNA near-diploid aneuploid sublines in ODFs and OPMLs appear as early events of the oral carcinogenesis in agreement with the concept of field effect. Near-diploid aneuploidization is likely to reflect mechanisms of loss of symmetry in the chromosome mitotic division. High DNA aneuploid and multiple sublines in OPMLs with dysplasia and OSCCs suggest, instead, mechanisms of “endoreduplication” of diploid and near-diploid aneuploid cells and chromosomal loss. High resolution DNA FCM seems to enable the separation of subsequent progression steps of the oral carcinogenesis.

Neuropathology of Olfactory Ensheathing Cell Transplantation into the Brain of Two Amyotrophic Lateral Sclerosis (ALS) Patients

Although a large number of amyotrophic lateral sclerosis (ALS) patients have undergone transplantation procedures with olfactory ensheathing cells (OECs) in the Bejing Hospital, to our knowledge, no post‐mortem neuropathologic analyses have been performed. We examined the post‐mortem brain of two Italian patients affected by ALS who underwent cellular transplantation in Beijing with their consent. Our aim was to assess the events following the graft procedure to possibly support the rationale of the treatment strategy. The neuropathologic findings were analyzed on the basis of the limited awareness of the experimental conditions and discussed in relation to the safety, efficacy and long‐term outcome of the transplanted cells. Islands of quiescent, undifferentiated cells within the delivery track persisting for up to 12 months–24 months were found. Prominent glial and inflammatory reaction around the delivery track strongly supports the encasement of the graft. Evidence of axonal regeneration, neuronal differentiation and myelination was not seen. The surgical procedure of implantation was not compatible with a neurotrophic effect. The OEC transplantation did not modify the neuropathology of ALS in the two patients. In conclusion, the present neuropathologic analysis does not support a beneficial effect of fetal OEC implantation into the frontal lobes of ALS patients.

Evidence for a possible anatomical subsite–mediated effect of tobacco in oral potentially malignant disorders and carcinoma

PURPOSE To test the hypothesis that cigarette smokers develop oral potentially malignant disorders or carcinomas in preferential anatomical subsites. METHODS The association of smoking habit with the presence of oral lesions in specific anatomical subsites was assessed in 123 patients using the odds ratio analysis. RESULTS When compared to all the other subsites, the relative frequency of smokers with lesions was higher in the buccal mucosa and in the floor of the mouth (FOM) (P=0.002 and P=0.005), while it was lower in the tongue (P<0.0005). Smokers were about 7 years younger than non-smokers (P=0.008). CONCLUSIONS The association of smoking and age suggests that smoking may contribute to generate a field of injury that leads to lesions in shorter periods than other causes. The stronger relationship of smoking with lesions in the buccal mucosa and FOM than in the tongue suggests that tissue characteristics mediate the effects of tobacco.

Clinical features of microinvasive stage I oral carcinoma

BACKGROUND This study aimed to analyse a case series of microinvasive (tumour thickness <4 mm) stage I oral squamous cell carcinoma (OSCC), with an emphasis on the clinical features of the tumours. METHODS In total, 32 microinvasive and 67 non-microinvasive stage I lesions, which had been surgically treated, were retrospectively studied and compared. The data analysed included gender, age, risk habits, clinical appearance, lesion site, symptoms, nodal involvement and outcome. RESULTS The clinical features of microinvasive lesions meant that, more often than not, they resembled premalignant lesions (P = 0.008), and diagnosis was mainly based on accurate clinical examination rather than the presence of symptoms (P = 0.029). During a median follow-up of 4.5 years, one nodal involvement and one cancer-related death were observed in patients with microinvasive lesions. A significantly higher (P = 0.044) level of nodal involvement was observed in the non-microinvasive lesion group. CONCLUSIONS   Stage I OSCC has a favourable prognosis overall, but nodal recurrence is more common in non-microinvasive cancers. As microinvasive lesions tend to present clinically as premalignant lesions, accurate clinical examination is essential if misdiagnosis of early lesions is to be avoided.