Comparison of different techniques for detection of Gal-GalNAc, an early marker of colonic neoplasia

The tumor marker, D-galactose-beta [1-3]-N-acetyl-D-galactosamine (Gal-GalNAc, also known as T-antigen) can be identified by a very simple galactose oxidase-Schiff's (GOS) reaction either on tissues or on rectal mucus samples from patients with colorectal neoplasms. Gal-GalNAc is expressed in the neoplastic mucosa as well as the remote non-neoplastic mucosa. It is, however, not expressed in colonic mucosa of normal subjects. We studied the expression of Gal-GalNAc by GOS reaction, lectin reactivity and immunocytochemistry in 10 normal, .45 precancerous [5 Crohn's disease, 15 ulcerative colitis (5 without dysplasia and 10 with dysplasia), 25 tubular adenomas], and 25 adenocarcinoma cases. Normal mucosa remote from tubular adenoma and adenocarcinoma was also studied. The GOS method was compared with reactivity of the lectin jacalin and immunostaining with antibody to T antigen (Anti-Tag Ab). GOS reaction was negative in all of the 10 normal specimens. Of the 5 Crohn's disease specimens, 2 were positive and 3 negative. In the 5 ulcerative colitis cases without dysplasia, positive reaction was seen in 2 cases and negative in 3. Of the 10 cases of ulcerative colitis with dysplasia, 5 showed positivity in dysplastic areas, and 3 of these were also positive in remote non dysplastic mucosa. Twenty of 25 tubular adenomas yielded a positive reaction in the adenoma, 14 of them showing positivity also in remote mucosa; 3 cases showed a positive reaction only in remote mucosa. Of the 25 adenocarcinomas, 21 showed a positive reaction in the adenocarcinoma as well as the remote mucosa. GOS reaction was intense in well differentiated adenocarcinoma and weak in poorly differentiated adenocarcinoma. Intense reaction was also seen in the intracellular mucus of some aberrant crypts and morphologically normal crypts remote from adenocarcinoma and tubular adenoma. GOS reaction showed an overall sensitivity of 75.7% and specificity of 100% for cancer and precancerous lesions. Jacalin reactivity was slightly more sensitive (84.3%) but less specific (80%) and Tag Ab reactivity even less sensitive (50%) but as specific (100%) for neoplastic and dysplastic mucosa. We conclude that the detection of the carbohydrate moiety Gal-GalNAc varies with the technique used. Compared to other techniques, GOS reaction is extremely simple and has a high degree of sensitivity and specificity. It can be used for detection of this tumor marker in remote non-neoplastic mucosa of patients with neoplasia or at risk of developing neoplasia. It, therefore, could be used as a cost effective screening test in rectal biopsy specimens of such patients.     

SNP genotyping using microsphere-linked PNA and flow cytometric detection.

BACKGROUND: Single nucleotide polymorphisms (SNPs) represent the most frequent form of genetic variations. Some of the most sensitive methods for SNP genotyping employ synthetic oligonucleotides, such as the peptide nucleic acid (PNA). We introduce a new method combining allele-specific hybridization, PNA technology, and flow cytometric detection. We tested the design by genotyping a Danish basal cell carcinoma cohort of 80 individuals for an A/C SNP in exon 6 of the XPD gene. METHODS: Genomic DNA was amplified by a two-step polymerase chain reaction (PCR) in the presence of fluorescein-dyed primers and fluorescein-12-dUTP. The allele-specific PNA molecules were covalently coupled to carboxylated microspheres with and without rhodamine. Allele-specific hybridization between PCR products and immobilized PNA was carried out at 60 degrees C followed by flow cytometric detection. RESULTS: We present a fully functional two-bead genotyping system based on PNA capture and flow cytometric detection used for the correct and fast regenotyping of a Danish basal cell carcinoma cohort. CONCLUSIONS: This new assay presents a simple, rapid, and robust method for SNP genotyping for laboratories equipped with a standard flow cytometer. Moreover, this system offers potential for multiplexing and will be operational for middle-scale genotyping.     

Tumor markers and lung cancer: correlation between serum and bronchial secretion levels of CEA, TPA, CanAg CA-50, NSE and ferritin

The levels of carcinoembryonic antigeny (CEA), tissue polypeptide antigeny (TPA), CanAg 50, neuron specific enolase (NSE) and ferritin were determined in bronchial secretion and serum of patients with neoplastic and non-neoplastic lung diseases. Simultaneous determination of two or three markers in the serum and in bronchoalveolar lavage (BAL) may be clinically useful for the diagnosis of lung cancer and even for the type of tumor. The positivity of CEA determined simultaneously in serum and in BAL of patients with lung cancer is higher than 80% whereas in patients with benign lung disease it is lower than 40%. The simultaneous assay of TPA in serum and in BAL showed 100% positivity in patients with oat-cell carcinoma, the frequencies of positivity were similar in patients with non-oat-cell carcinoma. For NSE and CanAg CA-50 patients with oat-cell carcinoma showed 100% positivity. Simultaneous assay of ferritin in serum and in BAL gave 85% positivity in patients with oat-cell carcinoma and only 23% in patients with non-oat-cell carcinoma. We conclude that the simultaneous determination of CEA and CanAg CA-50 or NSE in serum and in BAL is a useful aid in the diagnosis of lung malignancy.     

Immunogold study on lectin binding in the porcine zona pellucida and granulosa cells

An ultrastructural localization of lectin receptors on the zona pellucida (ZP) of porcine antral oocytes and on the granulosa cells was performed using a panel of horseradish peroxidase-labelled lectins in conjunction with antiperoxidase antibody and protein A-gold. In some cases, lectin incubation was preceded by sialidase digestion. WGA-, Con-A-, UEA-I-, RCA-I-, PNA- and SBA-reactive sites were distributed differently in the porcine ZP. Sialidase digestion increased the positivity obtained with RCA-I and it was necessary to promote PNA and SBA reactivity. These results indicated that the ZP contained N-acetylglucosamine, a-mannose, a-fucose, b-Gal-(1-4)GlcNAc, b-Gal- (1-3)GalNAc, b-GalNAc and sialic acid residues. We also observed the presence of vesicles in both the ooplasm and granulosa cells, showing a similar lectin binding pattern to that of the ZP, thus suggesting that the oocyte and granulosa cells are the site of synthesis of ZP glucidic determinants.     

Fine needle aspiration cytology in the diagnosis of bone lesions

OBJECTIVE: Fine needle aspiration cytology (FNAC) in combination with radiological examination has recently gained clinical recognition for evaluating skeletal lesions. We evaluated our experience with the use of FNA in diagnosing bone lesions with emphasis on areas of difficulty and limitations. MATERIALS AND METHODS: Over a period of 5 years FNA was performed in 66 cases of bone lesions. Aspirations were done by cytopathologists using 22-gauge needle. Out of 66 cases unsatisfactory aspirate was obtained in 12 cases. Cytohistological correlation was available in 19 cases. RESULTS: Adequate aspirates were categorized into neoplastic (27 cases) and non-neoplastic (27 cases) lesions. Of the 27 neoplastic aspirates, 20 were malignant (12 primary, 8 metastatic deposits) and 7 were benign. In the malignant group osteosarcoma was correctly diagnosed in 3 cases while other 3 were labeled as sarcoma NOS because of lack of osteoid. Metastatic deposits were sub-typed in 6 cases; from renal cell carcinoma (3 cases), proststic adenocarcinoma, follicular carcinoma thyroid, and squamous cell carcinoma. Neoplastic group comprised of 6 cases of cysts and 21 cases of chronic osteomyelitis. Thirteen cases were diagnosed as tuberculous osteomyelitis. CONCLUSIONS: FNA is a frequent indication in metastases in the bone where distinct cytologic features can even identify an unknown primary. However, diagnosis of primary tumours of the bone is limited by precise subtyping of the tumours. FNA has emerged as a cost effective tool for initial diagnosis of both neoplastic and non-neoplastic lesions of the bone.     

Changes in the glycosylation pattern during embryonic development of mouse kidney as revealed with lectin conjugates

Distribution of lectin-binding sites in adult and developing mouse kidney was studied with fluorochrome- and peroxidase-coupled lectins. Effects of fixation methods on lectin-binding patterns were also compared. Un-induced mesenchymal cells and ureter bud of the early metanephros reacted with Concanavalin A, Lens culinaris, Ricinus communis I, and wheat germ agglutinins, whereas binding sites for both soybean and peanut (PNA) agglutinins were seen only in ureter bud tissue. On induction, PNA positivity rapidly appeared in the induced, condensed areas of the metanephrogenic mesenchyme. Early glomeruli expressed heterogeneously terminal galactosyl and N-acetylgalactosaminyl moieties in the podocytes. Later, these sites disappeared and were apparently covered by sialic acids. Endothelia also displayed a comparable sialylation of terminal saccharide moieties during maturation. Binding sites for many of the above lectins were also found in the developing proximal and distal tubules. Terminal fucosyl residues, characteristic of mature proximal tubules, appeared during day 13 of development. Dolichos biflorus agglutinin reactivity, typically seen in the collecting ducts, appeared by day 13. Griffonia simplicifolia-I-B4 isolectin reactivity was exclusively localized to endothelial in adult kidney cortex, but in embryonic kidneys reactivity with collecting duct and podocytes was also seen. These results suggest that the compartmentalized expression of cell glycoconjugates in adult mouse kidney is acquired in a sequential manner during development. Such sequential appearance of the mature glycosylation pattern probably reflects functional maturation of the nephron.     

Histochemical demonstration of O-glycosidically linked, type 3 based ABH antigens in human pancreas using lectin staining and glycosidase digestion procedures

Histochemical analyses of the chemical structures of sugar sequences with or without blood group specificity were carried out by combined stepwise digestion of tissue sections with exo- and endoglycosidases and subsequent lectin stainings in formalin-fixed, paraffin-embedded human pancreas. In acinar cells from blood group A or AB secretor individuals, sequential digestion with alpha-N-acetylgalactosaminidase and alpha-L-fucosidase imparted reactivity with peanut agglutinin (PNA) in cells reactive with Dolichos biflorus agglutinin as well as those with Ulex europaeus agglutinin I(UEA-I). Simple fucosidase digestion imparted the PNA reactivity only in UEA-I reactive cells. Sequential digestion with alpha-galactosidase and fucosidase likewise liberated the PNA binding sites in Griffonia simplicifolia agglutinin I-B4 reactive cells from blood group B and AB secretors. Sialidase digestion liberated the PNA binding sites not only in acinar cells but also intercalated duct cells, islet cells of Langerhans and endothelial cells. The PNA reactivity obtained by these enzyme digestions was eliminted by endo-alpha-N-acetylgalactosaminidase (endo-GalNAcdase) digestion. Preexisting PNA affinity in acinar cells from non-secretors was also susceptible to endo-GalNAcdase treatment. Following the endo-GalNAcdase digestion, fucosidase or sialidase digestion recovered the PNA reactivity in acinar cells from nonsecretors. These results show that ABH determinants carried on O-glycosidically linked type 3 chain (D-galactose-(beta 1-3)-N-acetyl-D-galactosamine alpha 1-serine or threonine) are secreted in pancreatic acinar cells and suggest that product coded by the secretor gene is required for the complete conversion of type 3 precursor chains into H determinants.     

Detection of endometrial cancer by flow cytometry using two monoclonal antibodies.

BACKGROUND: To develop a supplementary diagnostic method for endometrial cancer by measuring the reactivity of various endometrial lesions with two monoclonal antibodies. METHODS: We investigated the reactivity of various endometrial lesions with two monoclonal antibodies (MSN-1 and MSN-3) by flow cytometry (one-color and two-color methods). RESULTS: The two-color method appeared to be suitable for use in place of simultaneous performance of the one-color methods with MSN-1 and MSN-3. The positivity rate for normal endometrium was 16.0% with the two-color method, which was lower than the rate of 30.0% obtained with concomitant used of the one-color methods. The positivity rate for endometrial cancer was high, 84.0%, with the two-color method. The positivity rate was 85.7% for well-differentiated endometrial cancer, 71.4% for moderately differentiated cancer, and 100.0% for poorly differentiated cancer; thus, the rate was high irrespective of the cellular differentiation. CONCLUSIONS: The two-color method is more useful than the one-color method as a supplementary diagnostic procedure for endometrial cancer.     

Localization of penultimate carbohydrate residues in zona pellucida and acrosomes by means of lectin cytochemistry and enzymatic treatments

Lectins from peanuts (PNA) and soy beans (SBA) bind terminal residues of galactose (Gal) and N-acetyl-galactosamine (GalNAc) respectively. Galactose oxidase oxidizes the hydroxyl group at C-6 of terminal Gal and GalNAc blocking the binding of PNA and SBA. Binding of these lectins to sugar residues is also severely limited by the existence of terminal residues of sialic acid. In the present study, lectin cytochemistry in combination with enzymatic treatments and quantitative analysis has been applied at light and electron microscopical levels to develop a simple methodology allowing the in situ discrimination between penultimate and terminal Gal/GalNAc residues. The areas selected for the demonstration of the method included rat zona pellucida and acrosomes of rat spermatids, which contain abundant glycoproteins with terminal Gal/GalNAc residues. Zona pellucida was labelled by LFA, PNA and SBA. After galactose oxidase treatment, terminal Gal/GalNAc residues are oxidized, and reactivity to PNA/SBA is abolished. The sequential application of galactose oxidase, neuraminidase and PNA/ SBA has the following effects: (i) oxidation of terminal Gal/GalNAc residues; (ii) elimination of terminal sialic acid residues rendering accessible to the lectins preterminal Gal/GalNAc residues; and (iii) binding of the lectins to the sugar residues. Acrosomes were reactive to PNA and SBA. No LFA reactivity was detected, thus indicating the absence of terminal sialic acid residues. Therefore, no labelling was observed after both galactose oxidase--PNA/SBA and galactose oxidase--neuraminidase--PNA/SBA sequences. In conclusion, the combined application of galactose oxidase, neuraminidase and PNA/SBA cytochemistry is a useful technique for the demonstration of penultimate carbohydrate residues with affinity for these lectins. This revised version was published online in November 2006 with corrections to the Cover Date.     

Utility of CD34 reactivity in evaluating focal nodular hepatocellular lesions sampled by fine needle aspiration biopsy

OBJECTIVE: To determine the patterns of CD34 reactivity in hepatocellular adenoma and focal nodular hyperplasia and to evaluate the utility of CD34 reactivity in the diagnosis of hepatocellular carcinoma. STUDY DESIGN: Seventeen cases of well-differentiated hepatocellular carcinoma, 14 cases of cirrhosis, 9 cases of focal nodular hyperplasia and 7 cases of hepatocellular adenoma were stained with immunoperoxidase antibodies to CD34. The slides were scored according to the degree of lesional reactivity. RESULTS: Fourteen of 17 cell blocks with hepatocellular carcinoma showed unequivocal sinusoidal or peripheral reactivity for CD34. Five of seven cases of hepatocellular adenoma and four of nine cases of focal nodular hyperplasia showed > 50% sinusoidal reactivity for CD34. All 14 cases of cirrhosis showed peripheral to no sinusoidal reactivity. CONCLUSION: CD34 reactivity in a diffuse sinusoidal pattern can be helpful in the diagnosis of hepatocellular carcinoma. However, consideration should be given to the possibility of hepatocellular adenoma and focal nodular hyperplasia, which can also exhibit significant diffuse CD34 reactivity. In these cases, a reticulin stain may be helpful with the differential diagnosis.     

Histochemical analysis of the chemical structure of blood group-related carbohydrate chains in serous cells of human submandibular glands using lectin staining and glycosidase digestion

Using lectin staining methods in combination with exo- and endo-glycosidase digestion procedures, we analyzed the chemical structure of different types of blood group-related substances in serous cells of formalin-fixed, paraffin-embedded human submandibular glands. Serous cells produced only H antigen; A and B antigens were not present, and the expression of H antigen is dependent on the secretor status of the tissue donor. Although reactivity with Ulex europaeus agglutinin I (UEA-I) was not markedly reduced by alpha-L-fucosidase digestion, an affinity for peanut agglutinin (PNA) was seen after fucosidase digestion in the cells from secretors. In those from nonsecretors, no PNA reactivity appeared after enzyme digestion. On the other hand, sialidase digestion elicited PNA reactivity in serous cells irrespective of the donor's secretor status. PNA reactivity observed after fucosidase or sialidase digestion was susceptible to endo-alpha-N-acetylgalactosaminidase (endo-GalNAc-dase) digestion. SBA reactivity in UEA-I-negative cells from secretors, or in cells from fetuses and newborn infants, was markedly reduced by beta-galactosidase digestion. After galactosidase digestion, reactivity with Griffonia simplicifolia agglutinin II (GSA-II) appeared in the corresponding cells. This GSA-II reactivity was almost completely eliminated by subsequent beta-N-acetylhexosaminidase digestion. Whereas PNA reactivity in these cells was not reduced by beta-galactosidase treatment, it was significantly diminished by endo-GalNAc-dase digestion. These results suggest that at least two kinds of precursor disaccharides are produced in submandibular serous cells, i.e., SBA-reactive D-galactose-(beta 1-3,4)-N-acetyl-D-glucosamine and PNA-reactive D-galactose-(beta 1-3)-N-acetyl-D-galactosamine alpha 1-serine or threonine (O-glycosidically linked Type 3 chain or T antigen). Final fucosylation and synthesis of these two types of precursor chain appear to be under the control of the secretor gene.     

胃粘膜肠上皮化生与胃癌关系的组织化学和免疫组织化学研究

应用ＡＢ（ｐＨ１．０）ＫＯＨ／ＰＡＳ粘液组织化学和ＡＢＣ法凝集素标记,对１９０例胃粘膜病变标本进行观察。结果表明,结肠不完全型肠上皮化生多见于肠型癌（ＩＴＣ）及其癌旁组织。两型肠化在弥漫型癌（ＤＴＣ）中无显著差异。５种凝集素受体的含量和分布的差异与胃癌的组织学类型和分化程度有关。ＷＧＡ、ＲＣＡ和ＰＮＡ主要标记在ＤＴＣ中,与ＩＴＣ相比,有非常显著的差异（Ｐ＜０．０１）。其染色水平随肿瘤分化程度的降低而升高。ＣｏｎＡ和ＤＢＡ主要标记在ＩＴＣ和伴有肠化的慢性萎缩性胃炎中。凝集素肠化分型与粘液肠化分型基本相符。我们认为结肠不完全肠化与ＩＴＣ的发生关系密切,而小肠型和结肠完全型肠化可能与ＤＴＣ的发生有关。     

A monoclonal antibody that binds to tumor-associated antigens of exfoliated cells in the smears of patients with cervical intraepithelial neoplasia

The reactivity of CE 407, a monoclonal antibody (MAb) known to bind to the cells of invasive cervical squamous cell carcinoma, was tested by the immunoperoxidase technique in samples from precursor lesions with and without associated condylomatous atypia. Antibody CE 407 bound with a high frequency to cells from cases of cervical intraepithelial neoplasia (CIN) and to cells showing condylomatous changes. Antibody CE 407 gave a positive reaction in 16 (64%) of 25 patients with CIN only. When the morphologic characteristics of human papillomavirus infection were present along with CIN, there was a higher frequency of positivity, with 27 (93%) of 29 such cases positive for CE 407. Reactivity of this MAb with normal exfoliated cervical epithelial cells was not observed.     

Vimentin-typing in diagnostic surgical pathology: a comparative study using four antibodies after different fixations

Vimentin-typing was carried out on various normal and neoplastic tissues using four anti-vimentin antibodies in order to evaluate the effect of different fixation treatments on tissue reactivity in comparison to the results obtained on frozen sections. All antisera were reactive on frozen material; on paraffin embedded material staining of tissues depended on the type of fixation method applied (formalin, methacarn or absolute alcohol) and each antibody behaved differently in relation to the fixative used. Only mesenchymal normal structures were revealed on frozen material whilst on paraffin embedded material three of the four antibodies reacted also with non-mesenchymal normal structures (epithelia, central and peripheral nervous system cells). All four antibodies decorated, regardless of treatment, neoplastic cells of mesenchymal and non-mesenchymal derivation, but not germ cells or germ cell tumors. The reactivity of vimentin to its specific antibodies depends on the fixative used: therefore, in routine pathology more than one antiserum should be available for testing. Furthermore, given the variety of non-mesenchymal structures stained by the anti-vimentin antibodies, the differential diagnosis of undifferentiated tumors must not be based on vimentin positivity alone. The expression of vimentin by non-mesenchymal neoplastic cells seems to parallel that of normal tissues during embryogenesis; therefore, this intermediate filament appears to be not only a marker of mesenchymal cells but also of many immature elements.     

Characteristics of the distribution of lectin receptors in intrahepatic cholangiocellular carcinoma

Summary The receptors of peanut agglutinin (PNA),Dolichos biflorus agglutinin (DBA) andUlex europaeus agglutinin I (UEA-I) were localized in intrahepatic cholangiocellular carcinoma, hepatocellular carcinoma, intrahepatic bile ducts and normal, cirrhotic and pericarcinomatous liver using the avidin—biotin—peroxidase complex method. It was found that epithelial cells of normal bile ducts had many UEA-I receptors, fewer DBA receptors and no PNA receptors. The positive rates of PNA, UEA-I and DBA receptors in 18 cases of intrahepatic cholangiocellular carcinoma were 88.9%, 61.1% and 33.3% respectively, which were significantly higher than those in hepatocellular carcinoma (16.0%, 4.0% and 4.0% respectively). Hepatocytes in normal, cirrhotic and pericarcinomatous liver had no receptors for these three lectins. It is suggested that lectin receptor distribution in intrahepatic cholangiocellular carcinoma is obviously different from that in normal bile duct cells and in hepatocellular carcinoma, and might be used as an auxiliary index in its clinical diagnosis.     

Adenoid cystic pattern in follicular variant of papillary thyroid carcinoma: a report of four cases

S. Mandal, and S. Jain
Adenoid cystic pattern in follicular variant of papillary thyroid carcinoma: a report of four cases Objective: An adenoid cystic pattern in thyroid tumours is a rare finding that may be seen in papillary carcinoma of thyroid (PCT), the follicular variant of PCT (FV‐PCT), a rare cribriform‐morular variant of papillary carcinoma of thyroid (CMV‐PCT) and follicular carcinoma. There is little published cytological literature describing these patterns. We report four cases of PCT with this unusual pattern. Methods: Fine needle aspiration (FNA) cytology was performed on four patients with a neck lump using a 22‐G needle; smears were stained with Giemsa and Papanicolaou stains. Immunocytochemical staining for thyroglobulin was done in all cases. Results: The patients were female and ranged in age from 18 to 46 years. They presented with a gradually increasing mass in the neck. FNA smears in all cases showed nuclear features of PCT. There were also prominent follicular areas with hyaline globules in some of the cell clusters reminiscent of adenoid cystic carcinoma and, in places, morula‐like groups of neoplastic cells were also seen. Immunocytochemistry for thyroglobulin was positive in all cases but negative in the hyaline globules. Conclusions: Adenoid cystic areas with morula‐like groups in PCT are a rare finding. Cytopathologists and clinicians should be aware of these distinct features in thyroid tumours to avoid diagnosing metastatic adenoid cystic carcinoma. It is also important to rule out CMV‐PCT since that variant is mostly associated with familial adenomatous polyposis, although sporadic occurrence is known.     

A first case of hepatocellular carcinoma in the baboon (Papio spp.) placenta

We present a case of hepatocellular carcinoma (HCC) in the placenta of healthy baboon (Papio spp.). Grossly, the fetal, maternal, and placental tissues were unremarkable. Histologically, the placenta contained an unencapsulated, poorly demarcated, infiltrative, solidly cellular neoplasm composed of cells that resembled hepatocytes. The neoplastic cells were diffusely positive for vimentin and focally positive for Ae1/Ae3, Arginase ‐1, glutamine synthetase, and CD10, and negative for ER, vascular markers (CD31 and D240), S100, glypican, C‐reactive protein, FABP, desmin, and beta‐catenin; INI1 positivity was similar to non‐neoplastic tissues. The case likely represents a unique subtype of HCC.     

Anti-Ki-67 peptide nucleic acid affects the proliferation and apoptosis of human renal carcinoma cells in vitro

We treated in vitro human renal carcinoma cells (cell line 786-0) with the lipid-delivered peptide nucleic acids (PNAs) against Ki-67 gene. Corresponding control groups were treated with the antisense oligonucleotides (ASOs) of the same nucleobase sequence, and with mismatched PNAs. In cells treated by anti-Ki-67 PNAs, the Ki-67 expression rate, Ki-67 protein level, cell growth and the DNA synthesis-indicative 3H-thymidine incorporation rate were lower than in the ASO-treated groups, and reduced significantly compared to untreated controls, whereas the rate of apoptosis was markedly increased by PNA treatment. We conclude that anti-Ki-67 PNA has more strong (than ASO) and dose-dependent effects on the proliferation and apoptosis of human renal carcinoma cells. Our results indicate that the strategy of using PNA against the Ki-67 gene might be a promising approach in renal carcinoma therapy.     

Bcl-2 protein in normal, hyperplastic and neoplastic breast tissues. A metabolite of the putative stem-cell subpopulation of the mammary gland

This investigation, though initially designed to examine the possible influence of the Bcl-2 protein on the node-metastasizing capacity of breast carcinomas, was amplified to study the expression of this anti-apoptotic protein in normal breast lobules and hyperplastic lesions. We examined paraffin sections of 508 breast carcinomas, stained for Bcl-2, estrogen (ER) and progesterone receptors (PgR) and epithelial membrane antigen, and occasionally for other antigens as well. Only a few cells showing a strong Bcl-2 positivity spotted the tubulo-lobular units of normal resting glands, whereas such cells were relatively numerous in atrophic lobules, and very scarce in the terminally differentiated lactating breast. Columnar and usual types of hyperplasia were exclusively, or almost exclusively, composed of Bcl-2(+), ER(+) and PgR(+) cells. The foci of carcinoma in situ and those of invasive carcinomas were respectively 83% and 66% positive for Bcl-2 in at least 25% of their cells. Even among the invasive carcinomas, Bcl-2(+) cases included 83% and 87% of the ER(+) and PgR(+) cases, respectively (p=0.0001). Though there was a statistically significant inverse relation between Bcl-2 and tumor grade (p=0.0001), no significant association was found between Bcl-2 and lymph node stage. In conclusion, we suggest that normal, hyperplastic and neoplastic breast epithelial cells expressing the anti-apoptotic protein Bcl-2 are immature cells that ought to form part of the stem-cell subpopulation, which is committed to the development and to the maintenance of the normal gland and which gives rise to hyperplastic and neoplastic disorders when its proliferation is deregulated. In ductal proliferative changes Bcl-2 assays may be useful for diagnostic but not for prognostic purposes.     

Immunohistochemical analysis of the ubiquitin-conjugating enzyme UbcH10 in lung cancer: a useful tool for diagnosis and therapy

The ubiquitin-conjugating enzyme (UbcH10) plays important roles in the regulation of cell cycle progression. Recently, UbcH10 expression has been demonstrated in several human and experimental tumors, and proteasome inhibitors have been tested in trials for pulmonary neoplasms; however, the underlying mechanisms as well as the clinicopathological relevance of UbcH10 in the genesis and progression of lung cancer remain largely unknown. Therefore, the authors evaluated the expression of UbcH10 in human lung cancer and evaluated its possible diagnostic and prognostic use. They found that most cases of lung adenocarcinoma, squamous cell carcinoma, and large cell and small cell carcinoma were positive for UbcH10. The expression levels of UbcH10 progressively increased with decreasing degree of tumor differentiation. There was a statistically significant difference of UbcH10 positivity between grade I/III of lung adenocarcinoma (p=0.013) and squamous cell carcinoma (p=0.002). No significant differences were found between histological types (p=0.072). In the case of cell blocks prepared from pleural effusions, inflammatory and reactive mesothelial elements did not show appreciable UbcH10 expression, whereas neoplastic cells exhibited clear UbcH10 positivity. The results suggest that UbcH10 might represent a new and promising diagnostic and prognostic marker in both histologic and cytologic specimens of lung cancer.