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PubMed2003

Suprabasal p53 immunostaining in premalignant endobronchial lesions in combination with histology is associated with bronchial cancer.

Endobronchial carcinoma develops through a continuum of morphologically recognizable pre-neoplastic changes. At present, no marker has been identified that can reliably predict the biological behavior of these lesions. Endobronchial lesions (n=39) sampled from patients (n=20) without clinically overt lung cancer, were analyzed by immunohistochemistry (IHC) for abnormal expression regarding the p53 protein, i.e. suprabasal p53 expression. Clear suprabasal p53 immunostaining was found in two (12%) of the hyperplastic or squamous metaplastic lesions, in one (10%) of the mildly or moderately dysplastic lesions and in nine (75%) of the severely dysplastic or carcinoma in situ (CIS) lesions. Suprabasal p53 immunostaining was found significantly more frequent in severe dysplasia or CIS (P<0.01). Of 17 patients follow-up data were available. After a median follow up of 7 months (range 2-37 months), six patients presented with bronchial carcinoma within the same lobe or bronchial spur where biopsies had been taken. Four of these patients revealed suprabasal p53 immunostaining in the biopsies obtained from the sites of future cancer. In three patients biopsies were obtained from future cancer sites as well as from distant sites in the ipsilateral lung. Suprabasal p53 immunostaining was found exclusively at future cancer sites of these patients (P=0.02). Suprabasal p53 immunostaining in addition to histology improved the specificity and the positive predictive value for bronchial carcinoma development in the same lobe or bronchial spur, compared with histology alone. These results suggest that suprabasal p53 immunostaining is associated with bronchial cancer and might have additive value to predict the biological behavior of pre-neoplastic endobronchial lesions in the population at risk of bronchial cancer.

Aged↗

PubMed1999

Subgroups of parkinsonian patients differentiated by peptidergic immunostaining of caudate nucleus biopsies.

In this study, Met-enkephalin (Met-enk), substance P (SP) and tyrosine hydroxylase (TH) immunostaining was assessed in caudate nucleus biopsies from 15 Parkinson's disease patients who were treated surgically. According to the combination of changes in Met-enk, SP and TH immunostaining, several subgroups of parkinsonian patients were disclosed. Group I: Patients showing low SP and normal Met-enk immunostaining, and variably reduced TH immunoreactivity. Group II: both SP and Met-enk immunostaining were apparently of normal intensity in these PD patients, but they showed the greatest decrease in TH labeling. Group III: PD patients that showed normal SP, very low Met-enk and variably reduced TH immunostaining. Low Met-enk immunostaining tended to correlate with the severity of the disease as judged by higher Unified Parkinson's disease Rating Scale and gait scores. These results suggest that different neurochemical phenotypes may exist among Parkinson's disease patients. Peptidergic deficits should be taken into account for therapeutic intervention.

Aged↗

PubMed2005

Loss of mismatch repair protein immunostaining in colorectal adenomas from patients with hereditary nonpolyposis colorectal cancer.

Colorectal adenomas occur at younger age, at increased frequency and have a greater tendency for malignant transformation in patients with hereditary nonpolyposis colorectal cancer (HNPCC). We performed immunostaining for the mismatch repair proteins MLH1, PMS2, MSH2 and MSH6 in 35 colorectal adenomas from 26 patients with HNPCC and identified loss of immunostaining in 23/35 (0.66) adenomas. Loss of mismatch repair protein immunostaining was particularly frequent in large (>5 mm) (14/16) and proximally located (13/15) adenomas, whereas the gene mutated--MLH1 or MSH2--and the type of mutation did not seem to affect the results. We conclude that loss of mismatch repair protein immunostaining is detected at a lower rate in adenomas than in carcinomas associated with HNPCC. Adenomatous tissue can thus be used for immunostaining of mismatch repair proteins in clinical investigations of HNPCC, but whereas loss of immunostaining may pinpoint the gene affected and thereby guide mutation analysis, retained staining cannot exclude that the adenoma developed as part of the syndrome due to reduced sensitivity. However, the analysis has a greater chance of being informative if large and proximally located adenomas are selected.

Adaptor Proteins, Signal Transducing↗

PubMed2002

The correlation among different immunostaining evaluation methods for the assessment of proliferative activity in uveal melanoma.

PURPOSE: Proliferative index of uveal melanoma cells serves as a prognostic factor. However, different methods are being used to determine proliferative index using immunostaining for proliferative markers. The major differences among assessment methods are whether the mean proliferative activity of all tumor cells in a section or for areas of rapidly proliferating cells is determined, and whether 10 or 20 fields are being evaluated. We aimed to assess the correlation among proliferative indexes obtained by different immunostaining evaluation methods. METHODS: Sections from 60 formalin-fixed paraffin-embedded uveal melanomas were immunostained with MIB-1 antibody. Immunostaining was assessed by counting immunoreactive cells in semi-randomly selected fields (non-selective method) and in areas with maximal immunoreactivity (selective method). Proliferative activity indexes according to the two methods in 10 and 20 high power fields (one high power filed = 0.785 mm(2)) were calculated and compared. RESULTS: The mean positive cell counts per mm(2) (MPCC/mm( 2)) according to the selective and the non-selective methods were 29.8 +/- 8.1 and 10.7 +/- 2.4 respectively (p = 0.004, paired t-test). Despite these different values, there was a good correlation between the MPCC/mm( 2) obtained by the non-selective and the selective methods for each tumor (r = 0.737, p < 0.0001, Pearson correlation). In addition, according to both methods, the readings of the first 10 fields and those of fields 1-20 correlated well. CONCLUSIONS: Mean proliferative activity of uveal melanoma cells correlates with the proliferative activity in localized areas of the tumor with rapidly proliferating cells. Therefore, uveal melanomas are classified similarly by the selective and non-selective methods of immunostaining evaluation. However, the two methods yield different proliferative index values for the same tumors, a fact that should be taken into account when comparing results of studies in which different techniques were used for immunostaining evaluation.

Antibodies, Antinuclear↗

PubMed2006

Mohs micrographic surgery for lentigo maligna and lentigo maligna melanoma using Mel-5 immunostaining: University of Minnesota experience.

BACKGROUND: Mohs micrographic surgery (MMS) continues to become a more common and accepted treatment for lentigo maligna (LM) and lentigo maligna melanoma (LMM). The primary difficulty encountered lies in the accurate identification of atypical single melanocytes to determine tumor-free margins. Numerous methods have been used to better visualize single melanocytes, with varying results. We present our experience using Mel-5 immunostaining in MMS of LM and LMM. METHODS: Two hundred patients with primary or recurrent LM or LMM were treated using MMS from 1999 to 2003 at the University of Minnesota. The initial clinical margins were determined by Wood's light examination, and an initial debulk specimen was taken and sent for formalin fixation and later reviewed by a dermatopathologist. The first Mohs layer was then taken, and staining with hemotoxylin and eosin as well as Mel-5 immunostaining was performed. All patients were followed up to evaluate for recurrence, with a mean follow-up time of 38.4 months. RESULTS: Of the 200 patients treated, only one recurrence was noted. This patient had been treated with excision followed by radiation before MMS. Use of Mel-5 immunostaining added approximately 40 minutes to each stage. Use of the Autostainer Immunostaining System (DAKO, Carpenterina, CA, USA) shortened the added time to 20 minutes. CONCLUSIONS: MMS with Mel-5 immunostaining yielded excellent results in the treatment of LM and LMM, with only one recurrence noted in 200 patients. When an automated immunostainer was used, minimal time was added to each Mohs stage.

Adult↗

PubMed2002

Cytokeratin and DAS-1 immunostaining reveal similarities among cardiac mucosa, CIM, and Barrett's esophagus.

OBJECTIVE: The normal histology at the gastroesophageal junction, and in particular the nature of cardiac mucosa, remains in dispute. Likewise, the relationship of intestinal metaplasia at the gastroesophageal junction (CIM) to Barrett's and intestinal metaplasia of the stomach (GIM) is unclear. The aim of this study was to assess the immunostaining characteristics of cardiac mucosa and CIM and compare their staining pattern with that of other foregut mucosal types. We hypothesized that the immunostaining patterns of these foregut tissues would provide insight into the nature and etiology of cardiac mucosa and CIM. METHODS: Paraffin-embedded biopsy specimens from 50 patients with normal antral or fundic mucosa, cardiac mucosa, squamous mucosa, CIM, GIM, or Barrett's were obtained and immunostained with a panel of monoclonal antibodies including those for cytokeratins 7 and 20 (CK7/CK20) and DAS-1. RESULTS: Biopsies from normal gastric antral and fundic mucosa and squamous esophageal mucosa all showed a non-Barrett's type CK7/CK20 immunostaining pattern, whereas in 85% of patients, cardiac mucosa had a Barrett's type CK7/CK20 pattern (p < 0.001). A Barrett's type CK7/ CK20 staining pattern was seen in 100% of Barrett's, 78% of CIM, and 0% of GIM patients. Likewise, DAS-1 staining was similar in patients with CIM and Barrett's and significantly different in patients with GIM. CONCLUSIONS: Cytokeratin immunostaining of cardiac mucosa demonstrates significant differences from recognized normal gastric and esophageal mucosa but a similarity to Barrett's. This suggests that cardiac mucosa, like Barrett's, may be acquired. Likewise, immunostaining similarities between CIM and Barrett's biopsies point to the possibility of a reflux etiology for CIM in some patients.

Antibodies↗

PubMed2005

Ultrarapid Ki-67 immunostaining in frozen section interpretation of gliomas.

BACKGROUND: Astrocytic tumours, the most common gliomas, are often classified intraoperatively using standard morphological staining. The final diagnosis and grading of gliomas on paraffin wax sections is often assisted by Ki-67 immunohistochemistry, but standard immunostaining protocols take too long to be used intraoperatively. AIMS: To investigate a new rapid Ki-67 immunohistochemical test for its use in an intraoperative setting. METHODS: The new Ki-67 immunostaining (Ultrarapid-Ki67) method on frozen sections can be carried out in 10 minutes. Thirty four pilocytic and diffuse astrocytomas were immunostained by rapid Ki-67 and results were compared with corresponding MIB-1 staining, histological grading, and prognosis. RESULTS: The staining protocol was practical to perform and the results were morphologically and quantitatively indistinguishable from those after immunostaining with MIB-1, an antibody recognising Ki-67 in paraffin wax embedded tissue. A comparison of Ultrarapid-Ki67 and MIB-1 immunostaining of paraffin wax sections showed almost identical quantitative correlation in astrocytic gliomas (r = 0.916; p<0.001). The Ultrarapid-Ki67 indices (percentage of positive cells) of low grade (I/II) astrocytomas ranged from 0% to 6.1%, whereas those of representative high grade (III/IV) tumours were significantly higher (range, 5.6-45%; p<0.001). The best prognostic cutoff point for Ultrarapid-Ki67 was 7.5%, which divided diffuse grade II-IV astrocytomas into significantly differing subsets (p = 0.0008). CONCLUSION: Ultrarapid-Ki67 immunostaining is a useful adjunct to morphological diagnosis and grading of astrocytic tumours, and as a fast test (approximately 10 minutes for staining plus three to four minutes for scoring), it could be used in routine intraoperative diagnosis of gliomas and other neoplastic diseases.

Adolescent↗

PubMed1988

Catecholaminergic innervation of luteinizing hormone-releasing hormone and glutamic acid decarboxylase immunopositive neurons in the rat medial preoptic area. An electron-microscopic double immunostaining and degeneration study.

Catecholaminergic innervation of luteinizing hormone-releasing hormone (LHRH) and glutamic acid decarboxylase (GAD) immunoreactive neurons in the rat medial preoptic area (MPO) was studied using electron-microscopic (EM) double-label immunostaining and combinations of single- and double-label immunostaining with acute axonal degeneration. The EM double-immunostaining experiments included double staining for either tyrosine hydroxylase (TH) and LHRH, or TH and GAD. Analysis of TH and LHRH double-immunostained material revealed synaptic connections between TH immunoreactive axons and LHRH immunopositive neurons. The TH and GAD double-staining experiments also demonstrated synaptic connections between axons immunoreactive for TH and GAD immunopositive neurons. Two days following unilateral surgical transection of the ventral and dorsal noradrenergic bundles, synaptic connections were found between degenerated boutons and GAD immunoreactive neurons in the ipsilateral MPO. However, no synapses could be observed in the same area between degenerated axons and the LHRH immunopositive neurons. Following the same operation and immunostaining for TH, a moderate number of degenerating TH axons as well as a large number of nondegenerated TH immunoreactive boutons were observed. Double immunostaining for TH and GAD in MPO sections ipsilateral to the operation revealed synaptic connections between the degenerating TH immunopositive axons and GAD immunoreactive neurons. These results suggest that there are direct synaptic connections between catecholaminergic axons and GAD and LHRH immunoreactive neurons in the medial preoptic area of the rat. Some of the connections between TH immunopositive afferents and GAD immunoreactive neurons may represent connections from noradrenergic neurons in the brain stem, while the majority of TH-GAD and TH-LHRH connections may represent innervation of GABA and LHRH neurons from local dopamine-containing cells.

Adrenergic Fibers↗

PubMed1996

MIB 1 immunostaining in cervical intraepithelial neoplasia: prognostic significance in mild and moderate lesions.

OBJECTIVE: MIB 1 is a new monoclonal antibody which recognizes nuclei of proliferating cells throughout the cell cycle except during the G0 and early G1 phases. In the present study we analyzed the MIB 1 immunostaining as an index of cellular proliferation in cervical intraepithelial neoplasia (CIN) and microinvasive carcinoma, with the aim to identify a relationship with the degree of dysplastic lesion and the risk of neoplastic progression. A correlation between the MIB 1 index and human papillomavirus (HPV) DNA presence was also investigated METHODS: Cervical bioptic samples were consecutively obtained from 86 women who attended our Colposcopic Service from January 1993 to June 1994, because of abnormal pap smears suspicious for cervical dysplasia and/or HPV infection. On histologic evaluation, 41 women had CIN, 23 cervical condyloma, and 22 squamous metaplasia. Ten patients with microinvasive squamous cervical carcinoma, matched for age and demographic characteristics, were selected from our series of cervical carcinomas and immunohistochemically analyzed. The expression of primary tumor cellular proliferation was immunohistochemically evaluated by monoclonal MIB 1 antibody (Immunotech, Marseille Cedex, France) on microwave oven-processed formalin-fixed paraffin-embedded tissue. Positive staining was expressed as the percentage of positive cells per 10(3) counted dysplastic cells for each case. RESULTS: A progressive significant increase in positive MIB 1 immunostaining was observed from squamous metaplasia to microinvasive carcinoma throughout the CIN lesions (p < 0.001). Considering only CINs, the MIB 1 index showed a significant increase with respect to CIN degrees (p < 0.001); no correlation was found between MIB 1 immunostaining and HPV infection, and lesion size. By analyzing the MIB 1 index with respect to CIN outcome in mild and moderate dysplasias, regressive lesions had lower values of MIB 1 immunostaining, while persistent and progressive lesions presented significantly higher positivity (p < 0.001). CONCLUSIONS: Our data demonstrated: (1) that positive MIB 1 immunostaining increased progressively from squamous metaplasia to CIN and microinvasive carcinoma, suggesting that neoplastic transformation is associated with a dysfunctional proliferation of cervical epithelium; (2) that there was a significant correlation between the MIB 1 index and CIN degree but not with respect to HPV DNA presence, and (3) that MIB 1 immunostaining might be useful for a clinical evaluation of mild and moderate dysplastic lesions. However, a much larger study needs to be done over a longer period of time to truly determine the value of the technique in prognostically predicting which lesions might or might not regress.

Adult↗

PubMed2002

A study of cytokeratin 20 immunostaining in the urothelium of neuropathic bladder of patients with spinal cord injury.

BACKGROUND: Normal urothelium is characterised by terminally differentiated superficial cells, which express cytokeratin 20 in the cytoplasm. In contrast, cultured human stratified urothelium, which does not undergo complete terminal differentiation of its superficial cells, does not express cytokeratin 20. If spinal cord injury (SCI) affects urothelial differentiation or induces squamous or other metaplastic change undetected by histological analysis, the superficial urothelial cells of the neuropathic bladder might be expected to show absence of immunostaining for cytokeratin 20. PATIENTS AND METHODS: We studied immunostaining for cytokeratin 20 in bladder biopsies taken from 63 consecutive SCI patients. Immunostaining was performed on paraffin-embedded tissue using a mouse monoclonal antibody (clone: Ks20.8). RESULTS: Of 63 biopsies, the epithelium was scarce in two. Eight biopsies showed squamous metaplasia and immunostaining for cytokeratin 20 was absent in all the eight biopsies. Of the remaining 53 cases, in which the umbrella cell layer of the urothelium was intact, immunostaining for cytokeratin 20 was seen only in ten biopsies. CONCLUSION: Superficial cells in the transitional epithelium showed immunostaining for cytokeratin 20 in 10 of 53 bladder biopsies taken from SCI patients. The reasons for this could be either that there is an underlying metaplasia or that changes in the neuropathic bladder affect urothelial differentiation. Taken with evidence from other systems, such as loss of cytokeratin 20 expression from static organ cultures of urothelial tissue, this might suggest that other factors, such as impairment of voluntary voiding in SCI patients, could affect expression of markers such as cytokeratin 20.

Adult↗

PubMed2006

Prevention of an additional surgery for regional lymphadenectomy in melanoma: rapid intraoperative immunostaining of sentinel lymph node imprint smears.

BACKGROUND: Sentinel lymph node (SLN) biopsy is performed at many institutions and is considered a standard of care in the management of cutaneous melanoma. The discriminatory immunostaining pattern with the 'MCW Melanoma Cocktail' (a mixture of MART-1 {1:500}, Melan- A {1:100}, and Tyrosinase {1:50} monoclonal antibodies) allows intraoperative immunocytochemical evaluation of imprint smears of SLNs for melanoma metastases. Cohesive cells of benign capsular melanocytic nevi that were also immunoreactive with the cocktail do not exfoliate easily for imprint smear detection. METHODS: We prospectively evaluated 73 lymph nodes (70 SLN & 3 non-SLN) from 41 cases (mean 1.8, 1 to 4 SLNs/case) of cutaneous melanoma using a rapid 17-minute immunostaining previously published protocol. The results were compared with permanent sections also immunostained with 'the cocktail'. RESULTS: 19.5%, 8/41 cases (12%, 9/73 lymph nodes) were positive for melanoma metastases on permanent sections immunostained with the 'MCW melanoma cocktail'. Melanoma metastases in 87.5% (7/8) of these cases were also detected in rapidly immunostained imprint smears, with 100% specificity and 90% sensitivity. None of the 7 SLNs from 7 cases with capsular nevi showed false positive results. CONCLUSION: Melanoma metastases could be detected in imprint smears immunostained with 'MCW Melanoma Cocktail' utilizing a rapid intraoperative protocol. The cohesive cells of the capsular nevi do not readily exfoliate and do not lead to false positive interpretation. In a majority of positive cases, a regional lymphadenectomy could have been completed during the same surgery for SLN biopsy and wide excision of primary melanoma site, potentially eliminating the need for an additional surgery.

Journal Article↗

PubMed1999

Lamina-specific synaptic activation causes domain-specific alterations in dendritic immunostaining for MAP2 and CAM kinase II.

Polyribosomal complexes are selectively localized beneath postsynaptic sites on neuronal dendrites; this localization suggests that the translation of the mRNAs that are present in dendrites may be regulated by synaptic activity. The present study tests this hypothesis by evaluating whether synaptic activation alters the immunostaining pattern for two proteins whose mRNAs are present in dendrites: the dendrite-specific cytoskeletal protein MAP2 and the alpha-subunit of CAMKII. High-frequency stimulation of the perforant path projections to the dentate gyrus, which terminate in a discrete band on the dendrites of dentate granule cells, produced a two-stage alteration in immunostaining for MAP2 in the dendritic laminae. Five minutes of stimulation (30 trains) caused a decrease in MAP2 immunostaining in the lamina in which the activated synapses terminate. After more prolonged periods of stimulation (1-2 hr), there was an increase in immunostaining in the sideband laminae just proximal and distal to the activated band of synapses. The same stimulation paradigm produced a modest increase in immunostaining for alpha-CAMKII in the activated laminae, with no detectable changes in the sideband laminae. The alterations in immunostaining for MAP2 were diminished, but not eliminated, by inhibiting protein synthesis; the increases in CAMKII were not. These findings reveal that patterned synaptic activity can produce domain-specific alterations in the molecular composition of dendrites; these alterations may be caused in part by local protein synthesis and in part by other mechanisms.

2-Amino-5-phosphonovalerate↗

PubMed1993

Detection of minimal residual disease by immunostaining of bone marrow biopsies after 2-chlorodeoxyadenosine for hairy cell leukemia.

2-Chlorodeoxyadenosine (2-CdA) yields high complete remission (CR) rates in patients with hairy cell leukemia (HCL). In an effort to detect minimal residual disease, we studied two B-lineage antibodies, L26 and MB2, and a T-lineage antibody, UCHL-1, in fixed marrow core biopsies from 34 patients with HCL before and after 2-CdA. Before therapy, hairy cells exhibited intense cytoplasmic membrane reactivity with L26 and strong intracytoplasmic reactivity with MB2. UCHL-1 did not react with hairy cells. Thirty-one patients were assessable 3 months after therapy. Five of 24 (21%) patients in CR by routine evaluation had residual HCL detected by immunostaining. Four of these 5 patients have been reevaluated at 1 year. One patient relapsed by routine evaluation, 2 remained positive by immunostaining alone, and 1 patient became negative by immunostaining. A total of 19 patients have been evaluated at 1 year. Only 1 additional patient has become positive by immunostaining alone. Immunostaining using the B-lineage antibodies highlighted the presence of hairy cells with preservation of morphology. This assisted in quantifying the extent of disease, particularly when hairy cells were interstitial and blended with surrounding hematopoietic tissue, when hairy cells were present in hypocellular marrows, when hairy cells were spindle-shaped, and when marrows were markedly fibrotic. Because immunostaining can be easily performed on routinely processed marrows, it is an attractive method to detect minimal residual disease. Our data suggest that some patients in apparent CR after 2-CdA may have minimal residual disease. Patients will need to be observed prospectively to determine if residual disease will be predictive of relapse.

Antibodies, Monoclonal↗

PubMed1990

Early diagnosis of cytomegalovirus hepatitis in liver transplant recipients: role of immunostaining, DNA hybridization and culture of hepatic tissue.

Immunostaining techniques that use a monoclonal antibody against an early cytomegalovirus antigen or a polyclonal antibody, in situ DNA hybridization and inoculation of cell cultures for the detection of cytomegalovirus from liver biopsy specimens were studied in 20 liver transplant patients with cytomegalovirus hepatitis, as defined by histological criteria. A total of 108 liver biopsy specimens from 20 patients with a diagnosis of cytomegalovirus hepatitis (obtained per protocol at 7, 21, 90, and 180 days or whenever liver dysfunction occurred), which had previously been examined histologically and in cell culture, were again studied by recutting the liver tissue for histological examination, DNA hybridization and immunostaining with monoclonal or polyclonal antibodies to cytomegalovirus. In 5 of 20 patients, the diagnosis of cytomegalovirus hepatitis could have been made earlier (mean = 9.6 days) by immunostaining with a monoclonal antibody. Of 47 biopsy specimens with cytomegalovirus inclusion bodies, the sensitivity and specificity of the diagnostic procedures were immunostaining with monoclonal antibody (84% and 90%) and polyclonal antibody (72% and 97%), in situ DNA hybridization (72% and 100%) and cell culture detection (52% and 95%), respectively. Immunostaining with a monoclonal antibody against an early CMV antigen frequently detected cytomegalovirus infection in the liver allograft earlier than identification of typical histological inclusion bodies. DNA in situ hybridization was less sensitive than other techniques but highly specific; cytomegalovirus cell culture lacked sensitivity compared with the other procedures.

Adult↗

PubMed1996

Microinvasive cervical carcinoma and cervical intraepithelial neoplasia: biologic significance and clinical implications of 72-kDa metalloproteinase immunostaining.

OBJECTIVE: The immunohistochemical expression of 72-kDa metalloproteinase was evaluated in cervical intraepithelial neoplasia (CIN) and microinvasive carcinoma, with the aim to define a relationship between 72-kDa metalloproteinase expression and neoplastic invasiveness, useful to identify subsets of intraepithelial lesions with higher risk of progression. MATERIALS AND METHODS: Cervical bioptic samples were obtained consecutively from 54 women who attended our Colposcopic Service from January 1993 to July 1993 because of abnormal pap smear, suspicious for cervical dysplasia and/or human papillomavirus infection. After written consent, 29 women with CIN were included in the study. All women with CIN 3 lesion underwent conization; in 21 women with mild or moderate cervical dysplasia, we did not perform any medical or physical treatment but followed them longitudinally at close interval. After 12 months, the clinical evolution was classified as spontaneous remission, persistence, or progression depending on the absence or presence of lesion and/or HPV infection in colposcopy, histology, and polymerase chain reaction findings. In the study we also included surgical specimens from 10 women with microinvasive squamous carcinoma who underwent primary radical surgery. Seventy-two kilodalton metalloproteinase positivity was immunohistochemically stained on serial sections by using the avidin-biotin complex technique (Vector Laboratories, Burlingame, CA) and expressed as percentage of cells per 10(3) counted neoplastic cells. RESULTS: Cytoplasmatic positive 72-kDa metalloproteinase immunostaining was significantly higher in microinvasive cervical carcinomas than in CIN lesion (Student's t test; P < 0.001). Considering only cervical intraepithelial neoplasias, a significant increase in 72-kDa metalloproteinase immunostaining was observed with CIN degree increasing (one-way analysis of variance; P = 0.002). No correlation was found between 72-kDa metalloproteinase immunostaining and HPV infection and lesion size defined by quadrants of the cervix involved with colposcopically evident dysplasia. By analyzing 72-kDa metalloproteinase positivity, regressive dysplasia showed low values of 72-kDa metalloproteinase immunostaining (median 1.2%, range 0.5-1.8%), while persistent (median 2.6%, range 1.9-3.6%) and progressive lesions (median 4.6%, range 2.3-6.9%) presented a significantly higher positivity (one-way analysis of variance; P < 0.001). DISCUSSION: In conclusion, the 72-kDa metalloproteinase expression is related to invasive potential with a significant increase in staining positivity in microinvasive carcinomas; 72-kDa metalloproteinase is detectable in cervical dysplasia, and it is related to the severity of cellular atypia. A clinical implication of 72-kDa metalloproteinase immunostaining seems to be indicated, by analyzing the differences in 72-kDa metalloproteinase positivity rates between regressive and persistent or progressive disease.

Adult↗

PubMed1997

Changes of cytoskeletal protein immunostaining in myelinated fibre tracts after focal cerebral ischaemia in the rat.

Breakdown of the cytoskeleton has been proposed to be a central event in the evolution of ischaemic brain damage. Alterations in the immunostaining of cytoskeletal proteins, particularly microtubule-associated protein (MAP) 2, have been suggested to be sensitive markers of ischaemic damage in the somatodendritic compartment. However, axons are also subjected to the adverse conditions created by an ischaemic challenge, but MAP2 is not located in the axonal compartment. The purpose of the present study was to examine immunostaining of beta-tubulin, MAP1a and MAP5, all of which are located in axons as well as perikarya, specifically in myelinated fibre tracts in rats subjected to unilateral middle cerebral artery (MCA) occlusion. In sham-operated control rats immunostaining of all three antibodies in myelinated fibre tracts had a smooth, regular appearance. Two hours after MCA occlusion there were striking changes in the patterns of immunostaining of all three antibodies in myelinated fibre tracts within the MCA territory. These were particularly noticeable in beta-tubulin- and MAP5-stained sections where the pattern in white matter had a rough, globular appearance. This pattern was accentuated at 6 h after MCA occlusion and the presence of "bulb-like" profiles in white matter tracts was notable particularly in the MAP5-stained sections. Thus, the changes in the patterns of staining at 2 h after MCA occlusion may represent the early stages of axonal disconnection, and immunostaining of microtubular proteins may represent a sensitive method to assess ischaemically induced damage to myelinated fibre tracts.

Animals↗

PubMed1998

Hamster pulmonary endocrine cells with positive immunostaining for calbindin-D28K.

We have examined the distribution of calcium-binding proteins (CaBPs) in adult and fetal lungs of Syrian golden hamsters (Mesocricetus auratus) using immunostaining with confocal laser microscopy and electron microscopy. Single and grouped (neuroepithelial body; NEB) endocrine cells were distributed from bronchi to alveolar ducts in the adult lung. Serial frozen sections immunostained for CaBPs in combination with immunostaining for endocrine markers such as calcitonin gene-related peptide, serotonin, PGP9.5, and synaptophysin revealed that positive immunostaining for calbindin-D28K (CB-D28K) was seen in single endocrine cells and NEBs. However, other so-called EF-hand family CaBPs, parvalbumin and calretinin, were not detected. Electron microscopically, positive immunoreaction for CB-D28K was mainly in the organelle-free cytoplasmic matrix of endocrine cells, and partly in nuclei and associated with secretory granules and endoplasmic reticulum. In fetal developing lungs, endocrine cells appeared first on gestational day 13, and they were positive for all the endocrine markers used. However, pulmonary endocrine cells were positively immunostained for CB-D28K from gestational days 15 and 16 onward. In summary, our observations suggest that CB-D28K is a useful marker for endocrine cells of the lung, and CB-D28K could function as a mediator of endocrine stimulation or calcium homeostasis in pulmonary endocrine cells.

Animals↗

PubMed2003

Microsatellite instability analysis and/or immunostaining for the diagnosis of hereditary nonpolyposis colorectal cancer?

Hereditary nonpolyposis colorectal cancer (HNPCC) represents 2-4% of colorectal cancers and is caused by a constitutional defect in a mismatch repair (MMR) gene, most commonly affecting the genes MLH1, MSH2, and MSH6. The MMR defect results in an increased cancer risk with the greatest lifetime risks for colorectal cancer and endometrial cancer. The HNPCC-associated tumor phenotype is generally characterized by microsatellite instability (MSI) and immunohistochemical loss of expression of the affected MMR protein. We have evaluated the information obtained from MSI analysis and immunostaining for MLH1, MSH2, and MSH6 in a series of 128 tumors from patients suspected of having HNPCC. A MSI-high pattern was present in 59 of 128 (46%) tumors. Loss of immunohistochemical expression for at least one of these MMR proteins was found in 54 of 59 (92%) evaluable MSI tumors. This loss affected MLH1 in 28, MSH2 in 22, and MSH6 in 21 tumors (with MSH6 as the only loss in 4 tumors). Five (8%) MSI-high tumors showed normal MMR protein expression. All 69 microsatellite stable or MSI-low tumors showed normal immunostaining for all three proteins. In 28 patients, all with MSI-H tumors, germ-line mutations of MLH1, MSH2, or MSH6 had been identified, and a corresponding immunohistochemical loss of MMR protein expression was identified in all these cases. In summary, immunostaining for the MMR proteins MLH1, MSH2, and MSH6 had a sensitivity of 92% and a specificity of 100% for detecting MMR-deficient tumors. MMR protein immunostaining facilitates mutation analysis in suspected HNPCC patients, since it pinpoints the mutated gene, but until the genetic background to the MSI tumors with retained MMR protein expression has been clarified, we suggest that MSI and MMR protein immunostaining should optimally be combined in clinical HNPCC analysis.

Adaptor Proteins, Signal Transducing↗