Cadaveric-biomechanical study on medial retinaculum: its stabilising role for the patella against lateral dislocation.

BACKGROUND: The aim of this study was to analyse the biomechanical role of medial retinaculum, as a stabilising factor against lateral patellar dislocation. MATERIALS AND METHODS: This cadaveric-biomechanical study included the patellae of 10 cadaveric knees, which were surgically exposed and the medial retinaculum of each one was located. A stable 24.51 N force was applied to the four parts of the quadriceps, and an increasing lateral displacing force was applied to the patella, up to 5 mm dislocation. The study was repeated for 0o, 45o, and 90o of knee flexion, with the medial retinaculum intact and dissected. The Wilcoxon signed rank test was used for data analysis. A p value < 0.05 was considered as statistical significant. RESULTS: After the dissection of medial retinaculum, the lateral displacement force was lower at every angle of knee flexion (p = 0.005, p = 0.007, p = 0.005, respectively). The lateral displacement force increased as the flexion angle increased (p = 0.005), regardless of medial retinaculum integrity. CONCLUSIONS: Medial retinaculum acts as a stabilising factor for the patella, against its lateral dislocation in lower flexion angles. Therefore, methods of surgical reinforcement or repair of medial retinaculum could provide protection against recurrent patellar dislocation.

Recombinative events of the T cell antigen receptor delta gene in peripheral T cell lymphomas.

Recombinative events of the T cell antigen receptor (TCR) delta-chain gene were studied in 37 cases of peripheral T cell lymphoma (PTCL) and related to their clinical presentation and the expression of the alpha beta or gamma delta heterodimers as determined by immunostaining of frozen tissue samples. There were 22 cases of alpha beta, 5 cases of gamma delta, and 10 cases of silent TCR expressing neither the alpha beta nor gamma delta TCR. 5 different probes were used to examine the delta locus. The 22 cases of alpha beta PTCL displayed biallelic and monoallelic deletions; a monoallelic V delta 1 J delta 1 rearrangement was observed in 1 case and a monoallelic germ line configuration in 7 cases. The 5 cases of gamma delta PTCL displayed biallelic rearrangements: the productive rearrangements could be ascribed to V delta 1J delta 1 joining in 3 cases and VJ delta 1 joining in 2 cases according to the combined pattern of DNA hybridization with the appropriate probes and of cell reactivity with the TCR delta-1, delta TCS-1, and anti-V delta 2 monoclonal antibodies. In the VJ delta 1 joining, the rearranged V segments were located between V delta 1 and V delta 2. Interestingly, in the third group of 10 cases of silent PTCL, 5 cases were found to have a TCR gene configuration identical to that in the TCR alpha beta PTCL, as demonstrated by biallelic delta gene deletion. These 5 cases were CD3 positive. The 5 remaining cases showed a monoallelic delta gene rearrangement with a monoallelic germ line configuration in 4 and a monoallelic deletion in 1. Four of these cases were CD3 negative, which was consistent with an immature genotype the TCR commitent of which could not be ascertained. Finally, TCR gamma delta PTCL consisted of a distinct clinical morphological and molecular entity whereas TCR alpha beta and silent PTCL had a similar presentation.

Assessment of vascular maturation in non-small cell lung cancer using a novel basement membrane component, LH39: correlation with p53 and angiogenic factor expression.

Angiogenesis, the formation of new vessels, has been demonstrated to be a potent and independent indicator of prognosis in non-small cell lung cancer patients. The extent of differentiation of the tumor vessels may affect access of peripheral white cells and egress or invasion of tumor cells. This has not been assessed in relation to tumor microvessel density or other variables and may be a marker of vascular remodeling. LH39 is a monoclonal antibody recognizing an epitope located at the lamina lucida of mature small veins and capillaries but not in newly formed vessels. We examined the ratio of mature:immature vessels in 81 non-small cell lung carcinomas and correlated the vascular maturation index (VMI) to different clinicopathological variables including angiogenesis. Mature vessels were defined by staining with antibodies to both LH39 and to CD31, using double immunohistochemistry, whereas immature vessels stained only for CD31. VMI was defined as the percentage fraction of mature vessels (LH39 positive)/total number of vessels (CD31 positive). The median VMI in lung carcinomas was 46% (range, 15-90%). There was a significant inverse correlation between high VMI and low thymidine phosphorylase expression (P = 0.0001), high VMI and nuclear p53 negativity (P = 0.01), high VMI and low angiogenesis (P = 0.0001), as well as between high VMI and absence of nodal involvement (P = 0.01). Low angiogenesis and high VMI were associated with a significantly better outcome (P = 0.0001 and P = 0.02, respectively). These findings show that there is a wide variation in the differentiation of tumor vasculature in lung carcinomas, and VMI gives new information on the degree of active tumor vascular remodeling independently from microvessel quantitation.

The expression of the epitope H recognized by the monoclonal antibody H is higher in astrocytomas compared to anaplastic astrocytomas and glioblastomas.

The epitope H contains an O-linked N-acetylglucosamine residue in a specific conformation and/or environment recognized by the monoclonal antibody H (mAbH). mAbH stains two bands with Mr x0.001 of 209 and 62 in lysates of cultured rat astrocytes. In normal human brains epitope H is absent from the overwhelming majority of normal astrocytes and only sparse reactivity is observed, confined mostly to fibrous astrocytes. Upregulation of the epitope H takes place in reactive astrocytes. In the present study we used the mAbH to investigate the immunohistochemical expression of the epitope H in 41 cases of astrocytic tumors including 19 cases of astrocytomas, 8 cases of anaplastic astrocytomas and 14 cases of glioblastomas. Seven out of 19 cases (37%) of astrocytomas showed weak staining, 10 cases (53%) moderate staining and 2 cases (10%) intense staining. Two out of 8 cases (25%) of anaplastic astrocytomas appeared negative, 3 cases (37.5%) showed weak staining and 3 cases (37.5%) moderate staining. Four out of 14 cases (28.5) of glioblastomas appeared negative, 7 cases (50%) showed weak staining, 2 cases (14%) showed moderate staining and only one case (7.5%) showed intense staining. There was a statistically significant elevation of the expression of the epitope H in astrocytomas compared to anaplastic astrocytomas and glioblastomas (p=0.047). These results indicate that the expression of the epitope H decreases in parallel with the increase of the grade of astrocytic tumors from low to higher grade neoplasms. This could be of interest for predicting the progression of an astrocytic tumor since it is documented that astrocytomas progress to tumors of higher grade of malignancy. Further investigation of the antigens bearing the epitope H might help to gain further insight into the mechanisms which regulate the progression of astrocytic tumors and to examine the relevance of the mAbH staining with respect to the prognosis of these neoplasms.

Epstein-Barr virus in T and natural killer (NK) cell non-Hodgkin's lymphomas.

Several studies using sensitive in situ hybridization techniques show that, in non-immunocompromised patients, Epstein-Barr virus (EBV) is more frequently detected in lymphomas expressing T cell markers than in B cell lymphomas. Among lymphomas expressing T cell markers, the presence of EBV is highly related to the site of origin of the tumor, being found in nearly all sinonasal lymphomas, in only a proportion of Waldeyer's ring, lung, gastrointestinal and nodal lymphomas, and undetectable in most primary cutaneous lymphomas. The role of EBV in their pathogenesis can be suggested in at least a proportion of extranodal lymphomas (nasal, lung, Waldeyer's ring, gastrointestinal) with T cell markers in which EBV genome is found in most if not all tumor cells (EBV-associated lymphomas) and the transforming LMP-1 protein is frequently expressed. Among these, sinonasal lymphomas constitute a distinct clinicopathologic entity which may present as lethal midline granuloma, are strongly associated with EBV and can be regarded in most cases as true NK cell lymphomas.

Middle ear adenoma. A tumor displaying mucinous and neuroendocrine differentiation.

Middle ear adenoma (MEA) is a distinctive, rare entity that appears to be derived from the lining epithelium of the middle ear mucosa. We report four cases of MEA displaying the typical histologic growth pattern. Two distinct tumor cell immunophenotypes were identified in all cases; the first type exhibited positivity with anti-epithelial membrane antigen and anti-keratin antibodies, and the second type showed immunoreactivity with anti-keratin, anti-vimentin, and anti-neuron-specific enolase antibodies. Ultrastructural studies revealed bidirectional mucinous and neuroendocrine differentiation, demonstrated by the presence of two distinct cell types containing apically located mucous granules and basally concentrated neuroendocrine granules, respectively. The presence of neuroendocrine differentiation was supported by the immunohistochemical detection of vasoactive intestinal polypeptide in the tumor cells in one case and neuron-specific enolase in three cases. These findings suggest that the potential for mixed mucinous/neuroendocrine differentiation described in other endodermally derived tumors also exists in middle ear mucosa. We also believe that the rare lesions diagnosed as primary carcinoid tumors of the middle ear might in fact be MEA with predominant or only neuroendocrine differentiation. The clinical course of our four cases and our review of the pertinent literature confirm the benign nature of MEA and indicate that these tumors should be treated by complete local excision without additional therapy.

A primary immunoblastic T malignant lymphoma of the small bowel, with azurophilic intracytoplasmic granules. A histologic, immunologic, and electron microscopy study.

We report an aggressive primary T-immunoblastic lymphoma of the small intestine without blood involvement or associated celiac disease. Grossly, the tumor was composed of multiple disseminated ulcerated, infiltrating, or protuberant nodular lesions. Immunologic investigation showed that lymphoma cells were of peripheral (post-thymic) T-cell origin and expressed the phenotype associated with cytotoxic-suppressor subset (Leu4/CD3+, Leu9/CD7+, Leu2/CD8+, Leu11/CD16+, Leu 7/NKcells+, FcIgG+, HLA-DR+, anti-Tac/CD25+, Ki-1/CD30-, Leu1/CD5-, Leu5/CD2-, Leu3/CD4-). A particular morphologic feature of this case is the presence of numerous azurophilic granules within the lymphoma cells, identified as lysosomes by cytochemical and ultrastructural studies. In view of recent immunologic evidence that normal cytotoxic/suppressor T-cells selectively reside within the epithelium of the normal bowel and some of them contain azurophilic granules, it could be suggested that our patient's lymphoma represents a malignant counterpart of these lymphocytes. Furthermore, the aggressive character of this T malignant lymphoma (T-ML) could be related to the expression of T-cell activation markers HLA-DR and Tac/CD25 and the proliferation-associated antigen Ki-67 on a high proportion of tumor cells.

Epstein-Barr virus (EBV) genomes and EBV-encoded latent membrane protein (LMP) in pulmonary lymphomas occurring in nonimmunocompromised patients.

Recently, in situ hybridization (ISH) techniques have shown that Epstein-Barr virus (EBV) could be detected in tumor cells of most angiocentric T-cell non-Hodgkin's lymphomas (NHL). These studies included only a few cases of T-NHL of the lung and pulmonary B-NHL and have not been investigated. Furthermore, the expression of the EBV-encoded latent membrane protein (LMP), which is known for its oncogenic properties, has not been reported. Twelve pulmonary NHL (six angiocentric T-NHL and six B-NHL) arising in nonimmunocompromised patients were examined for the presence of EBV-EBER mRNAs and LMP with ISH and immunohistochemistry, respectively. Four cases of pulmonary lymphomas arising in immunocompromised patients were also included in the study for comparison (one T-NHL in a patient under immunosuppressive treatment and three B-NHL in AIDS patients). EBV-RNA and LMP were detected in tumor cells in two of six nonimmunocompromised angiocentric T-NHL and in the four immunocompromised NHL. The six nonimmunocompromised B-NHL were EBV negative. These results suggest that EBV is associated with some angiocentric pulmonary T-NHL arising in patients without overt immunodeficiency whereas it is absent in such patients with B-NHL. The presence of the transforming EBV-encoded LMP in tumor cells suggests that EBV may be involved in the pathogenesis of some pulmonary T-NHL.

Expression of Epstein-Barr virus latent gene products and related cellular activation and adhesion molecules in Hodgkin's disease and non-Hodgkin's lymphomas arising in patients without overt pre-existing immunodeficiency.

Epstein-Barr virus (EBV) recently has been associated with Hodgkin's disease (HD) and the EBV genome was found in CD30-positive Reed-Sternberg cells. Therefore, tissue sections from 25 cases of HD, 35 cases of CD30-positive non-Hodgkin's lymphoma (NHL) (seven CD30-positive anaplastic large cell lymphomas [ALCLs] and 28 CD30-positive non-ALCLs), and 12 cases of CD30-negative NHL that previously had been screened for the presence of EBV by polymerase chain reaction and DNA in situ hybridization were studied by immunohistochemistry for the expression of the latent EBV proteins, latent membrane protein (LMP), and Epstein-Barr nuclear antigen-2 (EBNA-2). We also analyzed the expression of the B-cell activation molecule CD23 and the adhesion molecules LFA-1/CD11a and ICAM-1/CD54 because the upregulation of these molecules by LMP and/or EBNA-2 in vitro has been related to the EBV-induced lymphocyte growth. Latent membrane protein expression was found in Reed-Sternberg cells in nine of 25 cases (36%) of HD and in large, occasionally Reed-Sternberg-like tumor cells in six of 47 cases (12%) of NHL; these six tumors were CD30-positive, histologically high-grade NHL (one CD30-positive ALCL and five CD30-positive non-ALCLs). All the LMP-positive cases were also polymerase chain reaction EBV positive while LMP expression was not found in polymerase chain reaction EBV-negative HD and NHL. No staining for EBNA-2 was detected in our series. In view of the transforming potential of the LMP, these findings suggest that EBV may be associated with the development of some cases of HD and CD30-positive NHL. These findings also suggest a correlation between the expression of LMP and the detection of CD30 in tumor cells of HD and NHL. In contrast, no correlation was found between the expression of LMP and the detection of CD23, LFA-1/CD11a, and ICAM-1/CD54 in tumor cells of HD and NHL.

Bone marrow histologic and immunohistochemical findings in peripheral. T-cell lymphoma: A study of 38 cases.

The histologic and immunohistochemical findings in bone marrow (BM) biopsies from 38 patients with peripheral T-cell lymphoma (PTCL) are reported. Routine light microscopy showed that BM involvement was unequivocal in 12 cases and questionable in 14 cases. There was no histologic evidence of lymphoma in the remaining 12 cases. Immunohistochemistry performed on BM frozen sections demonstrated the T-cell origin of the infiltrating lymphoid cells in 24 of the 26 patients with unequivocal or questionable involvement. The malignant nature of these cells was suggested by demonstration of an aberrant T-cell phenotype identical to that observed in the other sites of involvement. In addition, in four of the 12 cases with apparently normal BM at routine light microscopy, immunohistochemistry revealed a minimal but phenotypically abnormal T-cell population, suggesting mild infiltration by lymphoma. These combined histologic and immunohistochemical data documented a high incidence (73%) of BM involvement by PTCL. In addition, a very peculiar sinusal pattern of BM involvement was found in five patients who presented an unusual type of hepatosplenic T-cell lymphoma expressing the gamma delta T-cell receptor. The present study demonstrates the high incidence of BM involvement by PTCL and emphasizes the value of frozen section immunohistochemistry to establish this diagnosis, especially when routine light microscopy findings are questionable.

Expression of p53 protein in T- and natural killer-cell lymphomas is associated with some clinicopathologic entities but rarely related to p53 mutations.

To determine if p53 abnormalities could be involved in the pathogenesis of T- or natural killer (NK)-cell lymphomas, we investigated 51 cases of these lymphomas for the expression of p53 and its relationship with p53 gene mutations, the expression of the p21 protein as well as the proliferative and apoptotic indices. Overexpression of p53 was found in 19 cases (37%), whereas mutations of the p53 gene were observed in only 5 of 28 tested cases. The analysis of immunohistochemical data showed some entity-related phenotypic profiles. Anaplastic large cell lymphomas showed a frequent overexpression of p53 (7/8 cases) and p21 (6/8 cases) proteins and rare p53 mutations (1/7 cases), suggesting accumulation of a functional wild type p53 protein able to induce p21 expression. Nodal peripheral T-cell lymphomas unspecified showed relatively frequent overexpression of p53 protein (5/7 cases), infrequent p21 expression (2/7 cases), and rare p53 gene mutations (1/6 cases). In angioimmunoblastic lymphomas, the common phenotype was p53-/p21- (15/17 cases), with only a few scattered p53-positive cells, which, on the basis of double staining results, were mostly Epstein-Barr virus-infected B cells. A p53 gene mutation was only found in 1 case (1/8 cases) of angioimmunoblastic lymphoma, which showed cytologic tumor progression. Mycosis fungoides showed p53 overexpression in 2 of 4 cases, including 1 case with p53 gene mutation and features of cytologic tumor progression. Nasal NK/T lymphomas showed p53 overexpression in 2 of 5 cases, 1 of which had a p53 gene mutation. Finally, all lymphoblastic T-cell lymphomas (5 cases) and gammadelta hepatosplenic T-cell lymphomas (3 cases) were negative for expression of p53 and p21 proteins. We conclude that p53 protein overexpression is a common finding in some entities of T- and T/NK-cell lymphomas, whereas a p53 gene mutation is a rare, sporadic, and rather late event associated with tumor progression in some instances. The p53/p21 expression pattern appears to be variable in T- and T/NK-cell lymphoma entities, reinforcing the concept of distinct, entity-related mechanisms of pathogenesis in these tumors.

Primary B-cell gastric lymphomas of mucosa-associated lymphoid tissue. Histological and immunohistochemical study of ten cases on surgical specimens.

Ten cases of gastric mucosa-associated lymphoid-tissue B-cell lymphoma were studied on surgical specimens by histology and immunohistochemistry, with monoclonal and polyclonal antibodies for B- and T-cells. For the first time, percentage of centroblast-like cells was appreciated, using their LN1 positivity, by opposition to the negativity of centrocyte-like cells (LN2 immunoreactivity alone). Lymphomas were divided into four main groups: A) centrocyte-like cells; B) centrocyte-like cells and immunocytoma; C) centrocyte-like cells admixed to centroblast-like cells; and D) centrocyte-like cells, immunocytoma and at least 30% of centroblast-like cells. Group C was divided into 3 subgroups: C1 (rare centroblast-like cells); C2 (30-50% of centroblast-like cells); and C3 (predominant centroblast-like cells). Therefore low grade (A, B, C1) and high grade malignancy groups (C2, C3, D) were identified: this preliminary subdivision could be extended in larger series and applied to mucosa-associated lymphoid-tissue lymphoma from other sites. Furthermore, the possible prognostic significance of this subdivision could be evaluated by correlation with long term follow-up.

Peripheral T-cell lymphoma or Hodgkin's disease in a HIV seropositive patient: a histopathological study.

Malignant lymphomas occurring in patients with AIDS are usually derived from the B-cell lineage while T-cell malignant lymphomas are very rare in these patients. We report a HIV seropositive 29-year-old homosexual man in whom cervical lymph node biopsy showed an atypical lymphoproliferative process. On morphological and paraffin section immunohistochemical grounds the possibility of Hodgkin's disease (HD) mixed cellularity was initially suggested, but frozen section immunohistochemical studies revealed that the cellular infiltrate exhibited an aberrant pan T immunophenotype and consequently the diagnosis of peripheral T-malignant lymphomas (T-ML) was made. However, genotypic studies would be required to definitely confirm this diagnosis, in such cases. In our case, varying numbers of small and medium-sized cells were positive for both Leu 3/CD4 and Leu 2/CD8 whereas some large cells reacted only with Leu 3/CD4 antibody. Some medium-sized, large and giant cells showed cytoplasmic positivity for Leu M1/CD15. Furthermore, the positivity of many large and giant cells with the activation markers BerH2/CD30, Ki-1/CD30, Tac/CD25 and HLA-DR suggested an activation state for these cells. Our findings emphasize the usefulness of frozen section immunohistochemical methods in order to investigate the spectrum of lymphoid malignancies occurring in HIV seropositive patients, and confirm results of previous studies which stressed the diagnostic difficulties that may appear in distinguishing HD from peripheral T-ML.

Serotonin-producing pancreatic endocrine tumour. Histological, ultrastructural and immunohistochemical study of a case.

Serotonin-producing pancreatic endocrine tumours are rare neoplasms which in most cases exhibit malignant biological behaviour. These tumours, in the majority of the well-documented cases, are composed of argyrophil- and argentaffin-positive cells which contain large pleomorphic neurosecretory granules. In contrast, argyrophilic non-argentaffin pancreatic endocrine tumours with tumour cells containing round neurosecretory granules are exceptional. In this study we describe such a tumour not associated with clinical evidence of carcinoid syndrome in a 60-year-old woman. Histological examination revealed tumour extension in pancreatic lymphatic vessels and veins but no evidence of locoregional or distant metastases. Ten months after surgery the patient showed no recurrence of the disease. Immunohistochemistry revealed cytoplasmic serotonin production in the tumour cells which were negative for anti-gastrin, insulin, glucagon, somatostatin, pancreatic polypeptide (PP), vasoactive intestinal peptide (VIP) and ACTH. This study emphasizes the usefulness of combined ultrastructural and immunohistochemical investigations in order to identify and characterize the rare pancreatic endocrine tumours with serotonin production.

Expression of p53, p21/waf1, bcl-2, bax, Rb and Ki67 proteins in Hodgkin's lymphomas.

The aim was to investigate the combined immunoexpression of p53, p21, bcl-2, bax, Rb and Ki67 proteins in Hodgkin's lymphomas (HL) and correlate expression patterns with the histotype and the Epstein-Barr Virus (EBV) status. Paraffin-sections from 56 cases of HL (18 nodular sclerosis and 38 mixed cellularity) and from ten "reactive" lymph nodes were investigated. P53, p21, bcl-2, bax, Rb and Ki67 proteins were detected in Hodgkin and Reed-Sternberg (HRS) cells in 35/56, 56/56, 24/56, 23/56, 56/56 and 56/56 cases of HL, respectively. No correlation was found between the expression of each protein and the EBV status or the histotype of HL. Comparison between p53 and p21 staining revealed two patterns: a) p53+/p21+ (35 cases); and b) p53-/p21+ (21 cases). The pattern p53+/p21+ suggests wild type p53 protein able to induce the expression of p21 while the p53-/p21+ pattern suggests p53-independent p21 expression. These results are consistent with the interpretation that inactivating p53 gene mutations may be rare in HL. Comparison between bcl-2 and bax staining showed a statistically significant relationship (p<0.001) for coexpression (19 cases) or absence of expression of both proteins (28 cases) in HRS cells. In contrast, bax expression was observed in most lymphoid cells in all "reactive" lymph nodes. Since the proapoptotic bax protein may act as tumour suppressor it is possible that the absence of this protein in HRS cells in a substantial proportion of HL may confer growth advantage and play a role in their pathogenesis. This could suggest bax gene alterations in some HL since in other studies acute lymphoblastic leukaemia cell lines demonstrate bax gene mutations with loss of bax immunoexpression. Another possibility is that reduced bax expression may be due to post transcriptional regulation, as was described in lymphoma cell lines. Comparison between Rb and Ki67 staining disclosed two main deviations from the normal parallel relationship in reactive lymph nodes: a) 2 cases with low Rb and high Ki67 expression possibly reflecting loss of Rb expression due to chromosome loss or to other abnormalities in the structure or the expression of Rb gene; and b) 9 cases with high RB and low Ki67 possible reflecting an attempt of Rb protein in excess to induce cell cycle arrest. Taken together, our findings provide combined immunohistological evidence for deregulated expression of cell-cycle and apoptosis-related proteins, that may play a role in the pathogenesis of HL.

Changing pattern of cytokeratin 7 and 20 expression from normal epithelium to intestinal metaplasia of the gastric mucosa and gastroesophageal junction.

It is currently unclear whether intestinal metaplasia at the esophagogastric junction and in the distal esophagus represent a continuum of the same underlying disease process, i.e., gastroesophageal reflux, or constitute different entities with a different pathogenesis. Biopsies below the Z line might show specialized epithelium in some patients and the question is whether this is another form of short segment Barrett's esophagus or whether it is related to a generalized atrophic process of the stomach. Data from recent studies regarding the expression of cytokeratin CK7 and CK20 in intestinal metaplasia (IM) found at the gastroesophageal junction are conflicting. Prompted by these data we undertook the present study: a) to evaluate the expression of CK7 and CK20 in IM of the gastric cardia and to compare the findings with those in patients with Barrett's esophagus and IM of the gastric corpus and antrum mucosa; and b) to evaluate the immunophenotype of non-intestinalized cardiac mucosa and to compare it with that of normal gastric epithelium. We studied the expression of CK7 and CK20 on biopsy specimens from patients with long-segment Barrett's esophagus (n=17) and surgical resection and biopsy specimens of gastric cardia (n=15), corpus (n=14) and antrum (n=22) from patients with histological evidence of IM. Eighty-four biopsy specimens from 42 patients (antrum n=15, corpus n=20, cardia n=7) without evidence of IM were studied as a control group. We observed an immunophenotype characterised by diffuse moderate to strong CK7 staining on the surface and crypt epithelium combined with strong CK20 staining on the surface and superficial part of the crypts in 94.1% (16/17) of the cases with long-segment Barrett's esophagus, but in none of the 36 cases with IM in distal stomach (antrum and corpus). IM in the gastric cardia expressed the immunophenotype seen in IM of the gastric mucosa in 93.3% (14/15) of the cases. On the other hand, normal cardiac epithelium expressed patchy strong CK7 staining on the surface epithelium and on both, superficial and deep parts of the pits combined with patchy strong CK20 staining on the surface epithelium and superficial pits, a feature permitting distinction of the normal cardiac epithelium from those of the normal gastric antrum and corpus epithelium. We conclude that the expression of cytokeratins 7 and 20 can be used to distinguish the origin of IM of the gastroesophageal junction. The CK7/20 immunophenotype of IM in the gastric cardia closely resembles that of the IM in the gastric antrum and corpus and is different from IM in long-segment Barrett's esophagus. In contrast, the CK7/20 immunophenotype of the cardiac epithelium is different from that of the gastric antrum and corpus mucosa, suggesting that cardiac epithelium might not be a native normal gastric epithelium but one that is acquired as a consequence of longstanding inflammation. Changing pattern of CK7 and CK20 expression from normal to intestinalized epithelium suggests that IM arising from cardiac epithelium might have distinctive features.

Tetranectin expression in gastric adenocarcinomas.

AIMS: The aim was to analyze the immunohistochemical localization of tetranectin in gastric adenocarcinomas and the adjacent tissues of the wall of the stomach. METHODS AND RESULTS: Forty cases of gastric adenocarcinomas were stained by the indirect immunoperoxidase method. Of the ten cases of mucinous signet ring cell carcinomas 5 showed high, 3 moderate and 2 low tetranectin expression. Of the ten cases of well-differentiated intestinal type adenocarcinomas (ITA) 4 showed moderate regional, 3 low regional and 3 negative tetranectin expression. Of the ten cases of moderately-differentiated ITA 3 showed moderate regional, 4 low regional and 3 negative tetranectin expression. Of the ten cases of poorly-differentiated ITA 4 showed focal low and 6 negative tetranectin expression. Overall, the mucinous signet ring carcinomas showed significantly higher tetranectin expression compared to ITA (chi2 = 3.95, p<0.05). In contrast, no significant relationship was found between tetranectin expression and the degree of differentiation in ITA (chi2 = 2.5, p>0.05). In all cases, the perineoplastic desmoplastic reactive stroma showed high expression of tetranectin intra- and extracellularly. The mast cells and goblet cells in the areas of intestinal metaplasia showed high tetranectin expression. CONCLUSIONS: This study shows that: a) tetranectin is produced and deposited extracellularly in the desmoplastic peritumoral stroma of infiltrating gastric adenocarcinomas; b) tetranectin is more highly expressed by the mucinous signet ring cell carcinomas compared to ITA; and c) the amount of tetranectin produced by the ITA is unrelated with the degree of tumor differentiation.

Immunohistochemical expression of p53, p21/waf1, rb, p16, cyclin D1, p27, Ki67, cyclin A, cyclin B1, bcl2, bax and bak proteins and apoptotic index in normal thymus.

The immunohistochemical expression of p53, p21, Rb, p16, cyclin D1, Ki67, cyclin A, cyclin B1, p27, bcl2, bax, and bak proteins and the apoptotic index (Al) were investigated in 20 normal thymuses (8 adults, 3 adolescents, 5 infants and 4 newborns). The expressions of Rb, Ki67, cyclin A and cyclin B1 were overlapping, being high in the cortex with a tendency for decreased expression toward the medulla. Apoptotic cells were mainly detected in the cortex and the corticomedullary junction, rarely being present in Hassall's corpuscles. The mean values of Ki67, cyclin A, and cyclin B1 expression in thymuses were 77.2%, 32.2% and 21.4% (newborns), 62.4%, 33.7% and 18.5% (infants), 56.9%, 23.4% and 18.9% (adolescents) and 38.7%, 21.7% and 14.6% (adults), respectively. The mean values of AI in thymuses from newborns, infants, adolescents and adults were 1.4%, 2.9%, 2.7% and 3.8%, respectively. This decrease in proliferation and increase in apoptosis may account for the process of thymic involution. P16 expression was widespread with most of Hassall's corpuscles being p16-positive. P16-positive cells and Hassall's corpuscles increased with the increase in age, in keeping with the suggested role of p16 in cellular senescence. P27 expression was undetectable in subcapsular thymocytes with a tendency for increased expression toward the medulla. The expressions of Ki67, cyclin A and cyclin B1 were inversly related with that of p27, consistent with previous evidence that p27 concentration is reduced when the cell-cycle progresses. P21 and much less frequently p53 proteins were mainly detected in a part of the subcapsular cortical epithelial cells. These findings suggest that a) in thymocytes, the apoptotic pathway is mostly p53-independent and the function of p21 as a negative regulator of the cell cycle must be redundant to other negative regulators, such as p16 and p27 which were abundantly detected in thymocytes and b) in some thymic epithelial cells, the p21 expression may be induced by p53, but in most of them seems to be p53-independent. Most of Hassall's corpuscles were p21-positive, consistent with previous evidence that these structures represent end stages of maturation of thymic medullary epithelium and that p21 protein is involved in the process of terminal differentiation. Cyclin D1 positivity was found in some macrophages. Bcl2 expression was mainly seen in medullary thymocytes, reflecting the surviving thymocytes in this region. The expressions of Bax and bak were more widespread in both the medulla and cortex, suggesting that these proteins play a broader role than bcl2 in the regulation of thymic apoptosis.

Immunohistochemical expression of the p53, mdm2, p21/Waf-1, Rb, p16, Ki67, cyclin D1, cyclin A and cyclin B1 proteins and apoptotic index in T-cell lymphomas.

Fifty-seven cases of T-cell lymphomas (TCL) including 5 lymphoblastic (T-LBL) and 52 peripheral TCL (PTCL) were analyzed by immunohistochemistry for the expression of p53, mdm2, p21, Rb, cyclin D1, cyclin A, cyclin B1, and Ki67/MIB1 proteins and 39/52 PTCL were also analyzed for the expression of p16 protein and for the presence of apoptotic cells by the TUNEL method. The aim was to search for abnormal immunoprofiles of p53 and Rb growth control pathways and to determine the proliferative activity and the apoptotic index of TCL. Abnormal overexpression of p53, p21 and mdm2, in comparison to normal lymph nodes, was found in 12/57, 10/57 and 2/57 cases of TCL, respectively. Abnormal loss of Rb and p16 expression was found in 1/57 and 2/39 cases, respectively, whereas abnormal overexpression of cyclin D1 was not detected in any of the 57 cases. Our data revealed entity-related p53/p21/mdm2 phenotypes. Indeed, most nodal and cutaneous CD30+ anaplastic large cell lymphomas (ALCL) showed concomitant overexpression of p53 and p21 proteins (7/8 cases), and mdm2 was overexpressed in 2 p53-positive nodal ALCL. In contrast, overexpression of p53 was found in 3/17 cases of nodal peripheral TCL unspecified (PTCL-UC) and 2/7 non-ALCL cutaneous pleomorphic TCL. Overexpression of p21 protein was detected in 2/3 p53-positive PTCL-UC and in 1/2 p53-positive non-ALCL cutaneous pleomorphic TCL. Finally, all the remaining 25 cases of TCL did not show p53 and p21 overexpression. Overall, the p53+/p21+ phenotype in 10/57 TCL suggests wild-type p53 capable of inducing p21 expression. The highest apoptotic index (AI) was found in ALCL and a positive correlation between apoptotic index and Ki67 index (p<0.001) was detected. Ki67, cyclin A and cyclin B1 expression was found in all 57 TCL and on the basis of the combined use of these 3 variables, 3 groups of proliferative activity could be determined: a) high in ALCL and T-LBL, b) low in mycosis fungoides (MF) and gammadelta hepatosplenic TCL, and c) intermediate in the remaining TCL entities. The proliferative activity in the 12 p53 overexpressing cases was higher in comparison to the 45 p53-negative cases. Ki67 expresion in more than 25% of tumour cells showed significant correlation with p53 overexpression (p<0.001). Rb expression tended to be parallel to Ki67, cyclin A and cyclin B1 expression in all but one case of nodal PTCL-UC which displayed loss of RB expression. Interestingly, this case was p53-negative, whereas the p53-positive cases were Rb-positive. These findings suggest that different pathogenetic routes may function in some TCL, involving either the p53 or, less frequently, the Rb pathways.

Expression of cyclin D3 and cyclin E and identification of distinct clusters of proliferation and apoptosis in diffuse large B-cell lymphomas.

In the present study 79 cases of de novo Diffuse Large B-cell Lymphomas (DLBCL) were studied in order: a) to analyse the expression of cyclin D3, cyclin E and cyclin D1 in relation to other proliferative features (expression of Ki67, cyclin A and cyclin B1), the apoptosis status and the expression of p53, Rb, p16 and p27; and b) to determine whether distinct clusters of proliferation and apoptosis could be identified in DLBCL. Overexpression of cyclin D3 and cyclin E was found in 35/79 (43%) and 18/79 (22%) cases, respectively, whereas overexpression of cyclin D1 was not detected in any case. In most cases (39/46) overexpression of cyclin D3 and cyclin E was mutually exclusive possibly reflecting different underlying pathways inducing deregulated expression of these cyclins. In most cases (29/35) overexpression of cyclin D3 was mutually exclusive with Rb/p16 aberrant expression status supporting an oncogenic role for cyclin D3 and suggesting that the pathogenetic effect of cyclin D3 overexpression occurs through perturbation of the Rb1 pathway. Combined alterations of the P53 and the Rb/p16/cyclin D3 expression status were significantly associated with higher mean values of cyclin A (p=0.023) and cyclin B1 (p=0.033) indicating that concurrent impairment of the p53 and Rb1 pathways induces increased tumour cell proliferation in DLBCL. Cluster analysis of the apoptosis and the proliferation status permitted separation of DLBCL into distinct groups with low (44 cases) and high (18 cases) apoptotic activity and into distinct groups with low (32 cases), intermediate (36 cases) and high (11 cases) proliferative activity. The identification of distinct clusters with respect to the proliferation and the apoptosis status indicates that groups with distinct cellular kinetic properties can be defined in the histological group of DLBCL.