Anti-cytokeratin Monoclonal Antibodies Research Articles

Prostatic perivascular epithelioid cell tumor: A rare entity and literature review

Perivascular epithelioid cell tumor (PEComa) is a rare mesenchymal neoplasm comprising perivascular epithelioid cells that express myomelanocytic immunophenotype, while stain negative for epithelial markers. We describe a case of prostatic PEComa in a 55-year-old man, who had one month history of frequent urination, hesitancy, and sensation of incomplete voiding. Radiological imaging disclosed prostatomegaly. Despite the medical treatment offered, episodes of urinary retention persisted. Subsequently, he underwent endoscopic transurethral resection of the prostate, histology showed tumor arranged in nests surrounded by thin delicate vessels. The tumor cells were epithelioid with abundant clear-to-eosinophilic cytoplasm, round nuclei, and inconspicuous nucleoli. Bizarre multinucleated giant cells, conspicuous mitosis and focal necrosis were evident. On immunohistochemistry, the tumor cells were diffusely positive for Human Melanoma Black (HMB45) and smooth muscle actin, negative for anti-cytokeratin monoclonal antibodies (AE1/AE3), Homeobox protein (NKX3.1), GATA Binding Protein 3 (GATA3), synaptophysin, Transcription Factor Binding to Immunoglobulin Heavy Constant Mu (IGHM) Enhancer 3 (TFE3), desmin, and SRY (sex determining region Y) -Box Transcription Factor 10 (SOX10). This uncommon case illustrates our diagnostic ordeal with a brief discussion on its nosology and a comprehensive literature review.

Mediastinal Extraskeletal Osteosarcoma in a Canine with Pulmonary and Cerebral Metastasis

Background: Extraskeletal osteosarcoma (EOS), a rare variant of osteosarcoma (OS), is a malignant neoplasm that develops in soft tissues without primary bone involvement. This study aims to describe a case of EOS with a mediastinal location in a canine.Case: A 10-year-old male, Uruguayan, Cimarron dog was presented to the Laboratório Regional de Diagnóstico, Faculdade de Veterinária, Universidade Federal de Pelotas (LRD/FV/UFPel) for necropsy. The dog had a history of submandibular swelling, progressive hind limb paralysis, muscle atrophy, and breathing difficulties. During necropsy, in the thoracic cavity, approximately 900 mL of serosanguinous exudate and a reddish-brown, bossed mediastinal mass measuring 15.0 cm in the longest axis were also noted. The lung exhibited multifocal to coalescent, white, firm nodules extending from the pleura to the parenchyma and measuring up to 4.5 cm in diameter. In the parietal and occipital region of the brain, a matte wine mass measuring 2.3 cm in the longest axis was observed. Fragments of the neoplastic mass, organs of the abdominal and thoracic cavities, and the brain were harvested and fixed in 10% buffered formalin. After 48 h, the samples were routinely processed, incorporated in paraffin, cut into 3 µm-thick sections, and stained using Hematoxylin and Eosin (HE). Selected sections of the neoplasm, the lung, and the brain were subjected to Von Kossa staining and immunohistochemical (IHC) analysis. For IHC, primary anti-cytokeratin monoclonal antibodies (clone AE1 / AE3, BioCare Medical) at a 1:100 dilution, vimentin (clone V9, BioCare Medical) at a 1:100 dilution, S100 Protein (clone 15E2E2, BioCare Medical) at a 1:100 dilution, and Ki67 (SP6 clone, BioCare Medical) at a 1:50 dilution were used. Immunostaining was visualized using 3-3' diaminabenzidine (DAB). Histological evaluation of the mediastinal mass, the pulmonary nodules, and the central nervous system among polygonal mesenchymal cells was conducted. Marked pleomorphism with euchromatic, rounded to oval nuclei, evident nucleoli, and poorly delimited eosinophilic cytoplasm. Neoplastic cells were arranged in nests and bundles with an invasive growth pattern. Osteoid and bone matrix formation as well as multinucleated giant cells of the osteoclast type were observed. The bone matrix was better evidenced in Von Kossa staining. IHC in all analyzed sections of the neoplastic cells showed positive immunostaining for Vimentin and Ki67. In the sections incubated with anti-cytokeratin and S100 protein antibodies showed no presence of neoplastic cells.Discussion: The diagnosis of EOS was based on the absence of primary bone lesions during microscopic necroscopy and on the exclusion of other histogenetic origins using IHC. The absence of primary bone lesions was the main attribute that differentiates EOS from other variants osteosarcomas, such as central/medullary and surface OS (periosteal and paraosteal) most frequently in canine species. The origin of EOS is uncertain. However, its occurrence has been originated with in pluripotent cells or previous injuries, such as retention of surgical sponges and vaccination sites. In this case, since the animal had no clinical history of injuries or surgical procedures that could induce the formation of a mediastinal neoplasm, the probable origin of the neoplasm was pluripotent cells. In dogs, EOS occurs mainly in the mammary glands, digestive system, liver, spleen, and subcutaneous tissue. Furthermore, the mediastinal location observed in this study was described only in goats. The clinical signs are nonspecific and varies with the location of the tumor, as observed in the present report. EOS may also present mediastinal location in the canine species. Necropsy, histopathological examination, and IHC were essential to establish the diagnosis of this OS variant.

Displasia renal em cães: estudo retrospectivo (2008-2013)

Background: Renal dysplasia, which has been reported in some dogs and human patients, refers to a developmental disorder of renal parenchyma due to imperfect inductive interaction between the mesonephric duct and the metanephric blastemal. In dogs, the characteristic histological findings on which diagnosis is based include (1) persistent metanephric ducts surrounded by primitive mesenchyme, (2) fetal or immature glomeruli, (3) fetal or immature tubules, and (4) anomalous presence of interstitial fibrous tissue. The aim of this study was to report the major pathological and immunohistochemical features of nine young dogs necropsied with renal dysplasia.Cases: The necropsy files from the Laboratório de Patologia Veterinária (LPV) of the Universidade Federal do Mato Grosso (UFMT) were reviewed between the years 2008 and 2013. Dogs diagnosed with chronic kidney failure and macroscopic and histopathological renal lesions consistent with renal dysplasia were selected. Kidney fragments in paraffin blocks were cut and stained with hematoxylin and eosin and by immunohistochemistry (IHC) using anti-vimentin and anti-cytokeratin monoclonal antibodies. The staining was considered positive for the presence of at least one renal cell marked with brown cytoplasmic staining clear and unambiguous. A total of 787 necropsies of dogs were performed. Of these, 64 had a clinical diagnosis of chronic renal failure of which 9 were classified as renal dysplasia. The age of the dogs ranged from 3 months to 2 years. Clinical signs were characterized by anorexia and non-regenerative anemia in 88.9%, vomiting 66.7%, dehydration 55.6%, uremia 55.6%, convulsion 33.4%, abdominal pain 22.3% and diarrhea in 11.2% of cases. At necropsy the main macroscopic changes in the kidneys were external surface with pale staining, in 6 of 9 dogs necropsied. Additionally, 5 dogs, cystic cavities of various sizes from 0.1 to 5 cm in diameter, diffusely distributed in the renal subcapsular surface and cutting were observed. Of the 64 dogs with diagnosis of chronic renal failure, 14.06% had dysplastic kidney changes, characterized by dilatation of Bowman’s space, glomerular and tubular atrophy, immature glomeruli and tubules, lymphocytic interstitial inflammation and fibrosis. In the dogs with renal dysplasia, it was observed that the tubular structures showed marked glomerular labeling for vimentin. Moreover, the kidneys of normal dogs showed weak or absent for marking tubes and glomerular structures for vimentin, and strong staining for cytokeratin in tubular cells, glomerular cells and collecting ducts.Discussion: Renal morphological damage observed in these nine dogs less than two years old, that contained degenerative and inflammatory changes, fibrosis and glomerular atrophy, but mostly immature glomeruli and tubules were associated with atypical renal dysplasia. This condition develops when the urethral diverticulum and metanephric blastoma not properly form in the embryonic stage, resulting in abnormal metanephric differentiation and formation of structures that do not recapitulate the normal nephrogenesis. Abnormal kidney function in young animals caused chronic renal failure resulting in death. The results of immunohistochemical observed in this study can complement the diagnosis of renal dysplasia. The morphological normality can be observed in the proportion of mesenchymal and epithelial tissue in the different structures of the kidney by staining with anti-cytokeratin and anti-vimentin. In the present study, there was moderate to strong labeling of vimentin in glomerular and tubular structures being dysplastic kidney would be expected to occur in normal dogs predominant staining with cytokeratin. Based on the clinic, pathological and immunohistochemical findings it is concluded that the animals developed renal dysplasia.

Nasal Transmissible Venereal Tumor In A Jack Russel Terrier Bitch

Abstract A two-year old Jack Russell Terrier female with left-sided epistaxis was brought to the Clinic of the Bucharest Faculty of Veterinary Medicine. Endoscopy of the left nasal cavity revealed a cauliflower-like mass. Cytological and histopathological features were specific to canine transmissible venereal tumour (CTVT). Tumor tissue showed positive immunoreactivity to anti-vimentin monoclonal antibody (90%) and Ki67 (35%) and samples were negative for anti-cytokeratin monoclonal antibody. Chemotherapy with vincristine sulphate at a dose of 0.025 mg/kg i.v./week for 6 administrations was successful. This paper presents a primary form of nasal CTVT in a Jack Russell Terrier female.

Predictive factors for the presence of tumor cells in bone marrow and peripheral blood in breast cancer patients.

Simultaneous detection of disseminated tumor cells (DTCs) and circulating tumor cells (CTCs) was shown to be associated with an especially poor prognosis and increased incidence of disease-related deaths in non-metastatic breast cancer patients. We analyzed the occurance of DTCs and CTCs in patients with primary breast cancer and evaluated the correlation of their presence with other prognostic markers and investigated the changes in DTCs/CTCs number at different time points during treatment.Blood of 50 patients with primary breast cancer were used for immunomagnetic separation and detection of circulating tumor cells using the commercial available system the AdnaTest Breast Cancer™ (AdnaGen GmbH, Langenhagen, Germany). Bone marrow aspirates from 50 patients were analyzed for DTCs by immunocytochemistry using the pan-cytokeratin antibody conjugated with FITC (Monoclonal Anti-Cytokeratin antibody F3418, Sigma Aldrich).DTCs were identified in 30% (15/50) and CTCs in 22% (11/50) of patients. We found that DTC positivity could point to asignificantly high risk of larger primary tumor size (p-value 0.011) and significantly higher risk of lymph node involvement (p-value 0.002). For CTC positivity, no such relationship was proven. DTCs have shown significantly higher prevalence in ER/PR-negative females and in HER2-positive cases. CTCs were equally prevalent in patients with the presence and absence of standard prognostic and predictive markers such as ER, PR and HER2. We found no correlation between CTCs and DTCs findings (r = -0.097, p= 0.504). We used DTCs/CTCs analysis for therapy monitoring in asmall group of 29 patients, who underwent neoadjuvant chemotherapy (NACT). We find out no significant correlation between DTCs/CTCs detection and the primary tumor response to NACT. Apathologic complete response (pCR) was achieved by 31% (9/29) of the patients in our study, however, no association was observed between pCR and the detection of DTCs after NACT.These results support the use of DTCs/CTCs analysis in early breast cancer to generate clinically useful prognostic information. The study of these cells apart from the impact on refining prognosis, has the exciting potential of individualising treatment for women with breast cancer. breast cancer, disseminated tumor cells, circulating tumor cells, bone marrow aspiration, prognostic/predictive markers, therapy monitoring.

Diffuse skin hyperpigmentation in CD30+ lymphoproliferation

CD30+ T-cell lymphoproliferative disorders (LD) comprise two main groups of diseases: CD30+ LD of the skin and systemic anaplastic large cell lymphoma (ALCL). The main feature of these disorders is the expression of CD30. We present a patient with an unusual clinical presentation of CD30+ lymphoproliferative disease in a 54-year old Caucasian male who presented with generalized lymphadenopathy and pronounced skin hyperpigmentation. In the lymph nodes and skin, CD30+ lymphoproliferation (ALCL) was diagnosed. The Prussian blue staining identified that the pigment responsible for the skin color was hemosiderin. Chemotherapy was started but the patient’s condition progressively worsened and he died a week after the first cycle. The complete color transformation of the entire skin due to hemosiderin accumulation is, to the best of our knowledge, the first reported observation in a CD30+ lymphoproliferation/ALCL patient. We speculate that hemosiderin-loaded macrophages resulted from the paraneoplastic process by some still unknown mechanism.

Accurate Detection of Carcinoma Cells by Use of a Cell Microarray Chip

BackgroundAccurate detection and analysis of circulating tumor cells plays an important role in the diagnosis and treatment of metastatic cancer treatment.Methods and FindingsA cell microarray chip was used to detect spiked carcinoma cells among leukocytes. The chip, with 20,944 microchambers (105 µm width and 50 µm depth), was made from polystyrene; and the formation of monolayers of leukocytes in the microchambers was observed. Cultured human T lymphoblastoid leukemia (CCRF-CEM) cells were used to examine the potential of the cell microarray chip for the detection of spiked carcinoma cells. A T lymphoblastoid leukemia suspension was dispersed on the chip surface, followed by 15 min standing to allow the leukocytes to settle down into the microchambers. Approximately 29 leukocytes were found in each microchamber when about 600,000 leukocytes in total were dispersed onto a cell microarray chip. Similarly, when leukocytes isolated from human whole blood were used, approximately 89 leukocytes entered each microchamber when about 1,800,000 leukocytes in total were placed onto the cell microarray chip. After washing the chip surface, PE-labeled anti-cytokeratin monoclonal antibody and APC-labeled anti-CD326 (EpCAM) monoclonal antibody solution were dispersed onto the chip surface and allowed to react for 15 min; and then a microarray scanner was employed to detect any fluorescence-positive cells within 20 min. In the experiments using spiked carcinoma cells (NCI-H1650, 0.01 to 0.0001%), accurate detection of carcinoma cells was achieved with PE-labeled anti-cytokeratin monoclonal antibody. Furthermore, verification of carcinoma cells in the microchambers was performed by double staining with the above monoclonal antibodies.ConclusionThe potential application of the cell microarray chip for the detection of CTCs was shown, thus demonstrating accurate detection by double staining for cytokeratin and EpCAM at the single carcinoma cell level.

Disseminated tumor cells in the bone marrow of patients with ductal carcinoma in situ

Detection of disseminated tumor cells (DTCs) in bone marrow is an independent prognostic factor in primary breast cancer. Here, we conducted a proof-of-principle study to evaluate whether this tumor cell spread occurs already in patients with ductal carcinoma in situ (DCIS). After preoperative screening by stereotactic core biopsy, 30 consecutive women with DCIS were included. Bone marrow aspirates, taken at the time of primary surgery, were subjected to DTC detection by a standardized immunoassay using the established monoclonal anti-cytokeratin antibodies A45-B/B3 and AE1/AE3. DTCs were detected in 4 of 19 cases of pure DCIS (21.1%) and in four of seven cases of DCIS with microinvasion (57.1%). After a median follow-up time of 22 months, two initially DTC-positive patients suffered from contralateral carcinoma and contralateral DCIS at months 12 and 30, respectively, whereas the remaining patients were relapse free. Thus, hematogenous tumor cell dissemination into bone marrow is an early event in breast cancer development.

Stem Cells Derived from Human Amniotic Fluid Contribute to Acute Kidney Injury Recovery

Stem cells isolated from human amniotic fluid are gaining attention with regard to their therapeutic potential. In this work, we investigated whether these cells contribute to tubular regeneration after experimental acute kidney injury. Cells expressing stem cell markers with multidifferentiative potential were isolated from human amniotic fluid. The regenerative potential of human amniotic fluid stem cells was compared with that of bone marrow-derived human mesenchymal stem cells. We found that the intravenous injection of 3.5 × 10(5) human amniotic fluid stem cells into nonimmune-competent mice with glycerol-induced acute kidney injury was followed by rapid normalization of renal function compared with injection of mesenchymal stem cells. Both stem cell types showed enhanced tubular cell proliferation and reduced apoptosis. Mesenchymal stem cells were more efficient in inducing proliferation than amniotic fluid-derived stem cells, which, in contrast, were more antiapoptotic. Both cell types were found to accumulate within the peritubular capillaries and the interstitium, but amniotic fluid stem cells were more persistent than mesenchymal stem cells. In vitro experiments demonstrated that the two cell types produced different cytokines and growth factors, suggesting that a combination of different mediators is involved in their biological actions. These results suggest that the amniotic fluid-derived stem cells may improve renal regeneration in acute kidney injury, but they are not more effective than mesenchymal stem cells.

Cytokeratin staining for complete remission in rectal cancer after chemoradiation

Pathologic complete remission (CR) of rectal cancer after neoadjuvant chemoradiation therapy (CRT) is generally confirmed by routine hematoxylin and eosin (H&E) staining. The aim of this study was to identify residual rectal cancer cells in primary lesions of patients with pathologic CR by immunohistochemical staining for cytokeratin. The medical records of 358 rectal cancer patients who underwent neoadjuvant CRT prior to radical surgery between October 2002 and August 2007 were reviewed. The authors stained sections of resected specimens of 58 patients (15.9%; 42 males; mean age 54 years) who achieved pathologic CR (as determined originally by H&E staining) with H&E and performed immunohistochemistry (IHC) using monoclonal anti-cytokeratin antibody. These stained sections were reviewed for residual rectal cancer cells by a pathologist. Of the 58 patients that achieved CR by initial pathologic examinations, eight (13.8%) were found to contain tumor by cytokeratin IHC. H&E staining revealed that six of these were positive for cancer cells, but the remaining two were negative for residual rectal cancer cells. Through better identification of residual rectal cancer cells, cytokeratin IHC offers a means of improving staging accuracy and thus provides useful information for prognosis and treatment decisions for patients with rectal cancer who had a clinical CR after CRT.

Hepatic carcinosarcoma with heterogeneous carcinomatous and sarcomatous elements: report of a case and review of the literature.

We report a rare case of hepatic carcinosarcoma consisting of two carcinomatous components and four sarcomatous components. A 54-year-old Japanese man presented with sudden right upper abdominal pain. Computed tomography showed a hepatic tumor measuring 17cm in the greatest dimension and intractable ascites, suggesting rupture of hepatocellular carcinoma (HCC). Transcatheter arterial embolization was repeated at our hospital, resulting in stabilization of the patient's general condition. Right anterior and left median sectionectomies were performed. Histologically, the tumor consisted of carcinomatous and sarcomatous elements. The carcinomatous elements consisted of moderately differentiated HCC (positive for hepatocyte antibody, murine monoclonal anti-cytokeratin antibody, and alpha-fetoprotein and negative for S-100 and desmin) and adenocarcinoma (negative for cytokeratin 7 and positive for carcinoembryonic antigen). Sarcomatous elements consisted of undifferentiated spindle cell sarcoma, rhabdomyosarcoma with acidophil striated cells including striped cells (desmin-positive), chondrosarcoma with cartilage matrix (S-100-positive), and osteosarcoma with osteoid. The sarcomatous elements were negative for epithelial markers. The patient is alive at 34months after the operation with peritoneal dissemination. A total of 23 cases, including the present case, of hepatic carcinosarcoma have been reported in the English language literature. This is the third reported case of hepatic carcinosarcoma with heterogeneous carcinomatous and sarcomatous elements.

Quantitative analysis of epithelial cells in urine from men with and without urethritis: implications for studying epithelial: pathogen interactions in vivo

BackgroundEpithelial cells in first catch urine (FCU) specimens from 87 men with and without urethritis were quantified. Epithelial cells were broadly categorised into transitional and squamous populations using morphological characteristics and immunostaining with anti-pan leukocyte and anti-cytokeratin monoclonal antibodies.FindingsThe majority (77/87 = 89%) of samples contained both transitional (76/87 = 87%; range 1 × 104 – 6 × 105, median 6 × 104) and squamous (57/87 = 66%; range 1 × 104 – 8 × 105, median 2 × 104) epithelial cells. The number of transitional cells correlated with the number of squamous cells (Spearman's rho = 0.697 p < 0.001). Squamous, but not transitional, cell numbers correlated with leukocyte numbers (Spearman's rho = 0.216 p = 0.045 and rho = 0.171 and p = 0.113, respectively). However there was no significant difference in epithelial cell numbers between men with and without urethritis. Nevertheless, some men with urethritis had relatively high numbers of transitional cells in their FCU. Transitional cells were morphologically heterogeneous and appeared to display complex cytokeratin phenotypes.ConclusionFurther studies are required to explore the complexity of epithelial cell populations in urine. These would provide novel opportunities for studying cellular interactions of C. trachomatis in male urethral infections, about which little is currently known.

The clinical significance of tumor cells in bone marrow or apheresis product and the efficacy of CD34+ selection and high‐dose chemotherapy in patients with Stage III breast cancer

The purpose of this study is to determine the presence of disseminated tumor cells in bone marrow or apheresis product, and also to evaluate the clinical significance of contaminated products and the efficacy of CD34(+) selection and high-dose chemotherapy in patients with Stage III breast cancer. Fifty-five patients were enrolled in this prospective cohort study. Whereas CD34(+) positive selection was not carried out in the first group (unselected group, n:31), CD34(+) positive selection was performed in the second group (CD34 selected group, n:24). Tumor cells were detected with anticytokeratin monoclonal antibody in the bone marrow, apheresis product and positive fraction. Tumor cells were found in six (19.3%) patients in unselected group and four patients (16.6%) in CD34 selected group (P = 0.76). The percentages of distant metastases were found higher in unselected group (51.6% vs. 25%, P < 0.01). Although there were no differences between the two groups for disease free survival (DFS; 44% vs. 74%, P = 0.24) or overall survival (54% vs. 68%, P = 0.84), DFS was significantly lower in patients with tumor cells than in patients without tumor cells (21% vs. 62%, P = 0.02). In conclusion, the presence of tumor cells in bone marrow or apheresis product decreases DFS in patients with Stage III breast cancer who underwent high-dose chemotherapy. CD34(+) selection does not change survivals, but it may decrease the distant metastases.

Monoclonal antibody to cytokeratin VKIALEVEIATY sequence motif reduces plasminogen activation in breast tumour cells

Cytokeratins (CKs) are the main structural proteins of epithelial cells. Although they mainly form cytoplasmic structures, they are also localized at the plasma membrane or secreted from the cells. Some CKs are over-expressed in tumour cells and are used as diagnostic and prognostic biomarkers. A stable hybridoma cell line producing anti-cytokeratin monoclonal antibody (anti-CK MAb) was prepared after immunizing mice with breast cancer MCF-7 cell lysate. As shown by 2D electrophoresis, immunoblotting and mass spectroscopy, the monoclonal antibody recognizes an epitope on CK1, CK2, CK8, CK10 and CK18 in MCF-7 cells. To identify the binding site of the antibody three peptides of 12 amino acids were synthesized, each overlapping a 27 amino acid consensus sequence of the recognized CKs. Anti-CK MAb expressed high affinity for a dodecapeptide with the sequence VKIALEVEIATY, localized in the CK α-helical B2 domain, as shown by ELISA and surface plasmon resonance. Treatment of MCF-7 cells by anti-CK MAb impaired plasminogen activation and consequently invasiveness of the cells. Our results show that, besides their use in diagnosis, anti-cytokeratin antibodies could be used in therapy of invasive breast cancer.

Bitumen products alter bax, bcl‐2 and cytokeratin expression: An in vivo study of chronically exposed road pavers

The skin of road pavers is chronically exposed to bitumen fumes, a mixture of volatile compounds and particles, containing several carcinogenic and non-carcinogenic polycyclic aromatic hydrocarbons. Bitumen fumes can alter cutaneous barrier integrity in various ways and induce skin diseases. The present study was devised to investigate the expression of apoptosis proteins (bax and bcl-2) and the cytokeratin pattern in skin specimens from road paving workers exposed to bitumen fumes using immunohistochemical techniques. Skin forearm punch biopsies from 16 occupationally exposed workers and an unexposed control group were processed for immunohistochemistry using a broad-spectrum anti-cytokeratin antibody and monoclonal antibodies for bax and bcl-2 immunostaining. Urinary 1-hydroxypyrene (1-OHP) was also determined. Morphological specimen evaluation showed epidermal thinning of exposed skin and flattened dermal papillae. In sections from exposed subjects, grade 3 bax overexpression and cytokeratin immunoreaction was detected in all layers, while bcl-2 expression was downregulated (grade 1) and confined to the basal layer. There was a significant difference in 1-OHP values between road pavers and the control group (p < 0.05). Overexpression of the cytokeratin pattern and bax and underexpression of bcl-2 in chronically bitumen-exposed skin suggest that bitumen fumes induce activation of apoptosis as a defense mechanism.

Epithelial microchimerism: Consistent finding in human liver transplants

Eleven liver biopsies from six male patients who received a liver transplant (LT) from female donors were examined in order to determine whether male host-derived hepatic cells were present in female grafts that exhibited minimal or important inflammatory damage. Immunohistochemistry for epithelial cell type differentiation (anticytokeratin monoclonal antibody) and fluorescence in situ hybridization for XY chromosomes identification were performed on each slide. Host-derived hepatic cells were found in all except one transplant, with a frequency ranging from 2.3 to 25 per thousand of the total hepatocytes in the biopsy specimen. They were usually found as isolated cells scattered throughout the hepatic lobule; in one patient they were grouped into little clusters. Host-derived hepatic cells persisted throughout the histological follow up (up to 535 days after LT). Polyploidy for XY chromosome was observed. Hepatocytes derived from extra-hepatic stem cells are frequently found in small numbers in human liver grafts and persist over time.

Ex vivo sentinel lymph node mapping in colon cancer: improving the accuracy of pathologic staging?

Background A subset of patients with colon cancer staged by conventional methods have occult micrometastases and do not receive adjuvant chemotherapy. Sentinel lymph node (SLN) mapping and staining by immunohistochemistry is a technique that may identify such occult micrometastases, thereby upstaging patients with positive findings. The purpose of this study was to determine whether ex vivo SLN mapping in colon cancer could be applied successfully to patients at our institution. Methods Seventeen patients with intraperitoneal colon tumors undergoing resection were studied prospectively. SLNs were identified as the first blue stained node(s) after ex vivo peritumoral injection of isosulfan blue dye. Additional lymph nodes were harvested and processed in accordance with standard pathologic evaluation for colon cancer. All nodes were examined after routine hematoxylin and eosin (H&E) staining. SLNs that were negative on H&E were analyzed further by multilevel sectioning and immunohistochemistry staining using anticytokeratin monoclonal antibody. Results Of the 17 study patients, SLNs were identified in 16 (94%) cases. The SLN was the only positive node in 3 patients. An identified SLN was positive (by H&E) in all patients with associated positive non-SLN nodes. The average number of nodes retrieved per patient was 16 (range, 4–54). Overall, SLNs accurately reflected the status of the entire lymph node basin in 16 (94%) patients. Two (12%) patients with negative nodes by H&E potentially were upstaged after further SLN analysis. The negative predictive value for SLN mapping was 89%. Conclusions The ex vivo technique of SLN mapping for colon cancer is feasible. In the current study, SLN results were concordant with non-SLNs in the majority of patients. Furthermore, this technique may have upstaged 2 (12%) patients. Whether this ultimately will affect overall survival has yet to be determined.

Specific Detection of Cytokeratin 20-Positive Cells in Blood of Colorectal and Breast Cancer Patients by a High Sensitivity Real-Time Reverse Transcriptase-Polymerase Chain Reaction Method

A real-time reverse transcriptase-polymerase chain reaction (RT-PCR) method for detection of cytokeratin 20-positive cells in blood characterized by two novel features was developed and tested on 99 patients with colorectal cancer, 110 with breast cancer, and 150 healthy subjects. To optimize the specificity and sensitivity of the method, two novel features were used. First, a primer overlapping two adjacent exons was generated to inhibit nonspecific amplification both in healthy donors and cancer patients; second, a non-end-point first-round amplification was used to increase sensitivity. The number of first-round cycles was chosen to reach the highest level of sensitivity while conserving quantitative characteristics. PCR efficiency increased from 88.9% in single-round RT-PCR to 99.0% in nested real-time RT-PCR. To establish sensitivity and specificity of the method, HT29 cells were serially diluted with normal blood. Detection limit improved from 100 HT29 cells (single-round RT-PCR) to 1 to 10 cells (nested real-time RT-PCR) per 3 ml of whole blood. None of the healthy subjects was positive, whereas 22 and 29% of all colorectal and breast cancer patients, respectively, had cytokeratin 20 cell equivalents in blood. The association between cytokeratin 20 cell equivalents and metastasis was statistically significant for breast (P = 0.026) but not colorectal cancer patients (P = 0.361). Negativity of all 150 healthy controls examined confers diagnostic potential to the method.

Immunohistochemical analysis of pelvic lymph node micrometastases in cervical cancer patients with histologically negative nodes

Objective：The purpose of this study was to identify, by immunohistochemistry, possible micrometastasis in the pelvic lymph nodes previously considered free by conventional histopathological examination, and to assess their influence on the survival of patients with cervical cancer. Methods：This retrospective study included patients (n=51) operated on between February 2001 and May 2004 for cervical cancer without histopathologic lymph node involvement. Lymph nodes (n=282) from 51 patients with histologically node-negative cervical cancer were evaluated for micrometastasis. These were submitted to immunohistochemical study using AE1/AE3 anti-cytokeratin monoclonal antibodies to identify neoplastic epithelial cells. Results：The mean age of the patients was 45.3 years (range, 27-72). Lymph node micrometastases were immunohistochemically detected in 3 of the 51 patients (5.9%), comprising 3 of 282 (1.1%) pelvic lymph nodes examined. One patient (Ib2) had adenocarcinoma and others (Ib1,Ib2), squamous cell carcinoma. All of them had negative lymph-vascular space invasion. In three patients, there were recurrences (66.7%, 2/3), and one patient (Ib1) died from the pelvic recurrence. Conclusion：We recommend an immunohistochemical examination for lymph node micrometastases in cervical cancer patients with histologically negative nodes. This immunohistochemical method can be employed successfully in the detection of neoplastic cells in lymph nodes previously considered free. Micrometastasis could provide important information for further treatment strategies and follow up. Its clinical significance in cervical cancer warrants further study.

Cytokeratins and Dermatology

Cytokeratins are fibrous intermediate-filament protein polymers present in almost all animal cells. Their function is related to epithelium structural maintenance, protection from mechanical trauma, and possibly communication between adjacent cells or cytoplasm components. Today there are 20 known cytokeratins, classified according to their molecular weight and pH as type I or acidic (cytokeratins 9-20) and type II or neutral-basic (cytokeratins 1-8). Cytokeratins are always expressed in specific pairs for each type of tissue, composed of one unit of type I and one unit of type II. Primary structural defects of cytokeratins are associated with various keratinization impairments. Two of the better characterized defects are bullous epidermolysis and epidermolytic hyperkeratosis. Anti-cytokeratin monoclonal antibodies are being used for diagnostic purposes to characterize the origin of poorly differentiated tumors and metastatic solid tumors.