Hormone receptor status is determined primarily to identify breast cancer patients who may benefit from hormonal therapy. The current clinical practice for the testing using either Allred score or H-score is still based on laborious manual counting and estimation of the amount and intensity of positively stained cancer cells in immunohistochemistry (IHC)-stained slides. This work integrates cell detection and classification workflow for breast carcinoma estrogen receptor (ER)-IHC-stained images and presents an automated evaluation system. The system first detects all cells within the specific regions and classifies them into negatively, weakly, moderately, and strongly stained, followed by Allred scoring for ER status evaluation. The generated Allred score relies heavily on accurate cell detection and classification and is compared against pathologists' manual estimation. Experiments on 40 whole-slide images show 82.5% agreement on hormonal treatment recommendation, which we believe could be further improved with an advanced learning model and enhancement to address the cases with 0% ER status. This promising system can automate the exhaustive exercise to provide fast and reliable assistance to pathologists and medical personnel. The system has the potential to improve the overall standards of prognostic reporting for cancer patients, benefiting pathologists, patients, and also the public at large.
Although all types of tumour and tumour-like conditions have been described to occur in the clavicle, they only contribute to less then 0.5% of all skeletal tumours. The incidence of primary chondrosarcoma of the clavicle is extremely rare. To our knowledge it has not been reported in Malaysia. We would like to highlight the possibility of chondrosarcoma as a differential diagnosis for a clavicular lesion.
Infantile myofibromatosis (IMF) is a rare tumour with a wide spectrum of disease activity ranging from a solitary cutaneous nodule through to a multicentric form with widespread visceral involvement. It is characterised by its unique ability to spontaneously regress and has a typical histological appearance of actin-positive fibroblasts arranged in whorls or fascicles and vessels in a pericytomatous pattern. A male infant with multiple lesions involving the subcutaneous tissue and bone from birth is described and followed-up for two years. Treatment of IMF is dependent on the location of the tumour/s with surgery or chemotherapy reserved for rapidly progressive or symptomatic disease. However, due to the low rate of recurrence and the possibility of spontaneous tumoral regression, therapeutic abstention, as practised in our patient, is justified.
The tumour suppressor genes, p53 and pRb, are known to play important roles in neoplastic transformation. While molecular routes to the uncontrolled growth of hepatocytes, leading to primary liver cancer have generated considerable interest, the roles of p53 and pRb mutations in hepatocellular carcinoma (HCC) and hepatoblastoma (HB) remain to be clarified. We examined the immunohistochemical expression of p53 and pRb gene products in 26 HCC and 9 HB, sampled into tissue microarray blocks. 10 (38%) of 26 HCC showed > 10% tumour nuclear staining for p53 protein, 3 of these also being HbsAg positive. Conversely, none of 9 HB expressed nuclear p53 immunopositivity. Some 24 (92%) HCC and 8 (89%) HB showed loss of pRb nuclear expression. Two of the 26 HCC and one of the 9 HB showed >10% tumour nuclear staining for pRb protein. Our results suggest that p53 does not have an important role in the development of HB but may contribute in HCC. There is also loss of pRb expression in the majority of HCC and HB, supporting loss of pRb gene function in the hepatocarcinogenesis pathway. However, a comparison of the staining profiles of p53 and pRb proteins in HCC and HB did not reveal a consistent pattern to differentiate between the two types of tumours immunohistochemically. Hence the use of p53 and pRB protein expression has no contribution in the situation where there is a diagnostic difficulty in deciding between HCC and HB.
Adjuvant! Online is a free web-based tool which predicts 10-year breast cancer outcomes and the efficacy of adjuvant therapy in patients with breast cancer. As its prognostic performance has only been validated in high income Caucasian populations, we validated the model in a middle income Asian setting.
A 35-year-old Chinese man who was known to have insulin-dependent diabetes mellitus was admitted for fever and weight loss. During his hospital stay, he fell to his death from his ward at the twelfth floor. The clinical features, radiological findings and gross organ changes at autopsy closely simulated miliary tuberculosis. Histology, however, revealed extensive necrosis of the adrenal glands, lungs, spleen, kidneys and thyroid associated with the presence of Histoplasma capsulatum organisms. This case highlights the similarity both clinically and pathologically between histoplasmosis and tuberculosis and emphasizes the need to be aware of this infection in a nonendemic area among patients with a compromised immune system.
Isosulfan blue is not available for clinical use in Malaysia. This study describes the use of methylene blue as an alternative to isosulfan blue in colorectal sentinel node mapping.
BACKGROUND: The interesting preponderance of Chinese with colorectal carcinoma (CRC) amongst the three major ethnic groups in Malaysia prompted a study to determine DNA mismatch repair (MMR) status in our CRC and attempt correlation with patient age, gender and ethnicity as well as location, grade, histological type and stage of tumour. Histologically re-confirmed CRC, diagnosed between 1st January 2005 and 31st December 2007 at the Department of Pathology, University of Malaya Medical Centre, were immunohistochemically stained with monoclonal antibodies to MMR proteins, MLH1, MSH2, MSH6 and PMS2 on the Ventana Benchmark XT autostainer. Of the 142 CRC cases entered into the study, there were 82 males and 60 females (M:F=1.4:1). Ethnically, 81 (57.0%) were Chinese, 32 (22.5%) Malays and 29 (20.4%) Indians. The patient ages ranged between 15-87 years (mean=62.4 years) with 21 cases <50-years and 121 ≥50-years of age. 14 (9.9%) CRC showed deficient MMR (dMMR). Concurrent loss of MLH1 and PMS2 occurred in 10, MSH2 and MSH6 in 2 with isolated loss of MSH6 in 1 and PMS2 in 1. dMMR was noted less frequently amongst the Chinese (6.2%) in comparison with their combined Malay and Indian counterparts (14.8%), and was associated with right sided and poorly differentiated tumours (p<0.05). 3 of the 5 (60.0%) dMMR CRC cases amongst the Chinese and 1 of 9 cases (11.1%) amongst the combined Malay and Indian group were <50-years of age. No significant association of dMMR was noted with patient age and gender, tumour stage or mucinous type.
The present study was conducted to assess utility of p16(INK4a) immunopositivity as a surrogate marker for genomic integration of high-risk human papillomavirus infection (hrHPV). A total of 29 formalin-fixed, paraffin-embedded cervical low-grade squamous intraepithelial lesions (LSILs), 27 high-grade squamous intraepithelial lesions (HSILs) and 53 invasive squamous cell carcinomas (SCCs), histologically-diagnosed between 1st January 2006 to 31st December 2008 at the University of Malaya Medical Centre were stained for p16(INK4a) (CINtec Histology Kit (REF 9511, mtm laboratories AG, Heidelberg, Germany). Immunopositvity was defined as diffuse staining of the squamous cell cytoplasm and or nucleus (involving > 75% of the intraepithelial lesions or SCCs). Staining of basal and parabasal layers of intraepithelial lesions was pre-requisite. One (3.4%) LSIL, 24 (88.9%) HSIL and 46 (86.8%) SCC were p16(INK4a) immunopositive. All normal squamous epithelium did not express p16(INK4). p16(INK4a) expression was significantly lower (p<0.05) in LSIL compared with HSIL and SCC with no difference in expression between HSIL and SCC.The increased p16(INK4a) immunopositivity in HSIL and SCC appears in line with the integrated existence of the hrHPV and may provide more insightful information on risk of malignant transformation of cervical squamous intraepithelial lesions than mere hrHPV detection.
Endocervicosis in the urinary bladder is a rare benign condition. We present a case in a 37-year-old woman with classical clinical and pathological features of endocervicosis. The unusual observation of endocervical-like mucinous epithelium in continuity with the urothelium in addition to fully developed endocervicosis prompted immunohistochemical profiling of the case using antibodies to cytokeratins (AE1/AE3, CK19, CK7, CK5/6, CK20), HBME-1, estrogen receptor (ER) and progesterone receptor (PR) to assess the relationship of the surface mucinous and endocervicosis glandular epithelia. The surface mucinous epithelium, urothelium and endocervicosis glands were immunopositive for AE1/AE3, CK7 and CK19 while CK20 was only expressed by few urothelial umbrella cells. The surface mucinous epithelium was CK5/6 and HBME-1 immunonegative but showed presence of ER and PR. This was in contrast to the urothelium's expression of CK5/6 but not ER and PR. In comparison, endocervicosis glands expressed HBME-1, unlike the surface mucinous epithelium. The endocervicosis epithelium also demonstrated the expected presence of ER and PR and CK5/6 immunonegativity. The slightly differing immunohistochemical phenotypes of the surface mucinous and morphologically similar endocervicosis glandular epithelium is interesting and requires further clarification to its actual nature. The patient has remained well and without evidence of disease 18-months following transurethral resection of the lesion.
On integration into the host cervical keratinocyte genome, human papillomavirus (HPV) E7 protein binds pRB,releasing E2F from normally incompetent pRB-E2F complexes and allowing propagation of G1-S transition by the E2F. p16(INK4a), a tumour suppressor protein, increases in reflex response to counter this. 29 histologically re-confirmed low-grade squamous intraepithelial lesions (LSIL), 27 high-grade squamous intraepithelial lesions (HSIL) and 30 invasive cervical squamous carcinoma (SCC) were immunohistochemically stained for p16(INK4a) expression using the CINtec Histology Kit (REF 9511, mtm laboratories AG, Heidelberg, Germany) to re-affirm the notion that integration of HPV occurs predominantly in SCC and possibly HSIL and less in LSIL and normal squamous epithelium (NSqE). Implicit was also the attempt to understand the role of E2F, as indicated by p16(INK4a), in evolution of SCC from HSIL. No ethnic predilection was noted for LSIL, HSIL or SCC. Patients with SCC were significantly older by about 14-years compared with HSIL (p < 0.05) while there was no significant age difference between HSIL and LSIL. p16(INK4a) expression was significantly increased (p < 0.05) in both HSIL (88.9%) and SCC (83.3%) compared with LSIL (3.4%) and NSqE (0%); the NSqE being normal squamous epithelium noted in 17 of the LSIL, 19 HSIL and 5 SCC. From these findings there is suggestion that fundamental upstream events viz HPV integration, E7 upregulation followed by E2F activation occurs at point of transformation to HSIL and continues unrelentingly for another one to two decades before hitherto unclear factors convert a non-invasive lesion into an overtly invasive malignant counterpart. Interestingly, the occurrence of HSIL and LSIL in almost the same age group could mean that alteration from episomal to integrated form of HPV may not incur a prolonged incubation period, unlike from HSIL to SCC.
With cervical carcinoma remaining the second leading cancer among Malaysian women, it is imperative to clarify the prevalence of human papillomavirus (HPV) in this respect, considering the dearth of local information.
Transaminase enzymes, alanine (ALT) and aspartate transaminase (AST), have been reported to be raised and implicated to have prognostic value in hepatocellular carcinoma (HCC). Ki67, a marker of cellular proliferative activity, has also been noted to be increased in HCC. A study was conducted at the Department of Pathology, Faculty of Medicine, University of Malaya, Kuala Lumpur to determine the possible association of proliferative activity, as determined by Ki67, with the transaminase enzymes. 31 cases of histologically diagnosed HCC who underwent tumour resection were retrieved from departmental archives. The patients' ages ranged between 40 to 79 years with a mean of 58.3 years. There was a male preponderance with M:F = 2.9:1. Ethnic Chinese formed 83.9% of the cases. 4 microm sections, cut from the formalin-fixed, paraffin-embedded tumour tissue block of each case, were immunohistochemically stained with Ki67 (DAKO monoclonal MIB-1) using the commercial DakoCytomation EnVision+System-HRP kit. The latest ALT and AST levels, assayed within 7 days prior to tumour resection, were retrieved from the patients' case records. 24 (77.4%) HCC demonstrated elevation of either ALT and/or AST. 27 (87.1%) HCC were immunopositive for Ki67. Ki67 immunoexpression was significantly correlated with raised transaminases (p<0.05). Hypothetically, the mechanism by which this phenomenon may occur may simply be release of transaminases due to destruction of hepatocytes by the cancer. Thus rising levels of the transaminases could signal a more rapid growth of the tumour and these routinely performed tests can be of prognostic value in management of HCC patients.
Cervical carcinoma, the second most common malignancy in Malaysian females, is aetiologically linked to the human papillomavirus (HPV). A study was conducted at the Department of Pathology, University of Malaya Medical Centre to compare the identification of HPV 6, 11, 16 and 18 in 40 archived formalin-fixed, paraffin-embedded cervical carcinoma by non-isotopic in-situ hybridisation (NISH) and polymerase chain reaction (PCR). HPV L1 ORF consensus PCR was also carried in cases which were negative on HPV type-specific PCR. NISH detected HPV 16 in 13 (32.5%) cases with one case demonstrating a concomitant HPV 18. beta-globin DNA PCR was carried out on the same paraffin block as for NISH in 27 cases and on a different paraffin block in 13, with amplification in 9 of the former and 3 of the latter. Thus only 12 cases were subjected to further HPV PCR. HPV was detected in 10 (83.3%) with HPV 16 in 9 cases and HPV L1 ORF in one. When using the same paraffin block for both methods of HPV detection, NISH detected HPV in 6 and PCR in 7. NISH failed to detect HPV in a case detected by PCR. 2 cases were negative for HPV using both methods. Hence, HPV detection results by NISH and PCR were concordant in 88.9%. Interestingly, NISH detected HPV in 2 cases with non-amplifiable beta-globin DNA. Using an alternative paraffin block for HPV PCR from NISH, HPV DNA was detected in 3 cases, two of which also showed type-specific positivity on NISH. The third case did not reveal type-specific positivity with NISH or PCR but demonstrated HPV DNA on L1 ORF consensus PCR. It thus appears that PCR and NISH can be successfully used to detect HPV in formalin-fixed, paraffin-embedded tissue and in the presence of intact DNA NISH may be as sensitive as PCR.
Sixteen low grade (LSIL), 22 high grade (HSIL) squamous intraepithelial lesions, 28 invasive (13 stage I and 15 stage II-IV) squamous cell carcinoma (SCC) and 15 benign cervices were immunohistochemically studied for involvement of Bcl-2 and Bax proteins in cervical carcinogenesis. 4-microm sections of the cases were immunostained for Bcl-2 (Clone 124: Dako) and Bax (Dako) and staining intensity was rated as 1 (light), 2 (moderate) and 3 (strong) and percentage cellular staining as 0 (negative), 1 (1-25%), 2 (26-50%), 3 (51-75%) and 4 (>75%) with score derived by multiplying staining intensity and percentage positivity. The cut-off value, indicating upregulated expression, was computed as >2 for Bcl-2 and >8 for Bax. Bcl-2 was upregulated (p < 0.05) in HSIL and Bax in SCC when compared with benign cervical squamous epithelium. Bcl-2 expression was confined to the lower third of the epithelium in the benign cervices and LSIL. In HSIL, expression reached the middle and upper thirds. 4 (30.8%) HSIL with upregulated Bcl-2 demonstrated intensification of staining around the basement membrane. SCCs showed "diffuse" (evenly distributed) or "basal" (intensified staining around the periphery of the invading tumour nests) expression of Bcl-2. Of the 5 SCCs with upregulated Bcl-2, 1 of 2 (50%) stage I and 3 (100%) stage II-IV tumours exhibited the "basal" pattern. Benign cervical squamous epithelium, LSIL, HSIL and SCC showed a generally diffuse Bax expression. Thus, Bcl-2 and Bax appeared to be upregulated at different stages of cervical carcinogenesis, Bcl-2 in HSIL and Bax after invasion. Intensification of staining of Bcl-2 at the basement membrane in some HSIL and SCC is interesting and may augur for increased aggressiveness.
Hepatocellular carcinoma (HCC) ranks as the fifth most common cancer with an increasing frequency worldwide. "Nuclear atypia", one of the critical features in histological diagnosis of malignancy and grading of the tumour, is generally ascertained through eyeballing. A study was conducted at the Department of Pathology, University of Malaya Medical Centre to assess whether nuclear area, (surrogate measure for nuclear size) and standard deviation (surrogate measure for nuclear pleomorphism) when objectively measured via computer-linked image analysis differs between (1) benign and malignant liver cells and (2) different grades of HCC. A 4-microm thick H&E stained section of 52 histologically re-confirmed HCC with 36 having benign, non-dysplastic surrounding liver were analysed using the Leica Q550 CW system. 10 consecutive non-overlapping, non-mitotic and non-apoptotic nuclei of HCC and surrounding benign hepatocytes respectively were manually traced at 400x magnification on the computer monitor and the nuclear area for the particular cell computed in arbitrary units by the Leica QWIN software. A total of 360 benign hepatocytic nuclei, 240 low grade HCC and 280 high grade HCC nuclei were traced. The mean nuclear area of the benign hepatocytes (37.3) was significantly smaller (p < 0.05) than that of both low grade (65.2) and high grade HCC (80.0). In addition, the mean nuclear area of high grade HCC was significantly larger (p < 0.05) than the low grade HCC. SD of the nuclear areas was lowest in benign hepatocytes (9.3), intermediate in low grade HCC (25.0) and highest in high grade HCC (25.6). These findings indicate that computer-linked nuclear measurement may be a useful adjunct in differentiating benign from malignant hepatocytes, in particular in small biopsies of well-differentiated tumours, and in predicting survival after surgical resection and transplant.
Ki-67 expression in diffuse proliferative lupus nephritis, WHO Class IV, was compared against normal controls to establish that cellular proliferation is involved in the production of glomerular hypercellularity. Twenty-three histologically confirmed WHO Class IV lupus nephritis and 23 normal control renal tissue were immunohistochemically stained with a polyclonal antibody to Ki-67 (Dako) using the peroxidase labelled streptavidin bioitin kit (Dako). There were 20 females and 3 males, with 17 Chinese and 6 Malays in the WHO Class IV lupus nephritis group. Ages of patients ranged between 10-56 years with a mean of 31.9 years. The normal controls, 20 males and 3 females, and ethnically 9 Indians, 7 Malays, 2 Chinese, and 5 foreign nationals (4 Indonesians and 1 Bangladeshi), had an age range between 15-33 years (mean = 23.3 years). Sixteen (69.6%) WHO Class IV lupus nephritis and 8 (34.8%) normal controls demonstrated Ki-67 immunoreactivity in at least 1 glomerulus (p<0.05). Of the 256 WHO Class IV lupus nephritis non-sclerosed, glomeruli studied, 37 (14.5%) were Ki-67 immunopositive compared with normal controls where 16 (0.7%) of 2159 glomeruli demonstrated Ki-67 (p< 0.01). Cellular proliferative activity, as evidenced by Ki-67 expression, was significantly increased in WHO Class IV lupus nephritis confirming that cell proliferation contributes to glomerular hypercellularity.
p53 is the most commonly mutated gene in human cancers. It encodes a 53 kilodalton protein with several evolutionarily conserved domains viz sequence-specific DNA binding, tetramerisation, SH3 molecule binding, C-terminal and N-terminal. Existing in the cell at a very low level and in a relatively inactive form, p53 protein is increased and activated during periods of cellular stress. Unlike other proteins, the increase in protein level and its activation result from modification of the protein rather than genetic transcriptional or translational upregulation. Normally, Mdm2 protein interacts with p53 protein and effectively targets it for ubiquitin proteolysis within an autoregulatory feedback loop. Phosphorylation at the N-terminus reduces p53 interaction with Mdm2 with a resultant increase in p53 protein level. Modification at the C and N termini via phosphorylation or acetylation upregulates binding to specific DNA targets increasing transcription of these downstream genes. The net effect of p53 protein increase and activation lies in arrest of the cell in cycle which allows time for repair of the incurred damage or apoptosis or death of the cell. Failure of these normal protective and adaptive mechanisms caused by mutation of the p53 gene with product of an abnormal protein, loss of p53 protein through interaction with and degradation by HPV E6 protein or overexpressed Mdm2 etc. permits DNA-damaged cells to continue replicating. Left unchecked, this frequently contributes to tumourigenesis. Various methods have been devised to screen for mutations of the p53 gene, still the most common source of failed p53 mechanism. These include immunohistochemical detection of mutated proteins or identification of altered electrophoretic mobility of mutated p53 sequences. Sequencing of the gene nonetheless remains the most accurate method for determination of mutation. Major advances have been made in p53 research but the most meaningful probably lies in the promising results achieved in tumour therapy where introduction of wild type p53 gene has resulted in regression of non-small-cell lung cancer (NSCLC). Many other notable developments in this field include description of p53 homologues, "gain of function" mutants, p53 polymorphisms, angiogenesis-inhibiting properties of wild type p53 protein etc.
A study was conducted at the Department of Pathology, University of Malaya Medical Centre, Kuala Lumpur into the histological type (WHO classification), grade (modified Edmondson and Steiner's grading system), mitotic rate, bile production, hyaline globule and Mallory hyaline formation of 52 cases of hepatocellular carcinoma (HCC) diagnosed during a 13-year period between 1st January 1990 to 31st December 2002. In addition, associated cirrhosis, dysplasia (large liver cell dysplasia: LLCD and small liver cell dysplasia: SLCD) and microvascular permeation were also looked for whenever the situation permitted. The patients' ages ranged from 21-years to 85-years (mean = 58.7 years) with a predilection for males and Chinese. Histologically, majority (73.1%) of the tumours demonstrated a trabecular pattern of growth. The bulk (73%) of the tumours were either of grade II or III differentiation. Mitotic activity ranged between 0-100/10 high power fields (hpf) with a mean of 22.2/10 hpf. Bile was noted in 25%, hyaline globules 17.3% and Mallory bodies in one case. Concomitant cirrhosis was present in 73.5%. 73.5% of the cases had associated LLCD. 5 with LLCD also showed SLCD. Microvascular permeation was shown in 76.2% of cases. On comparison with findings from other studies, no major difference seems to exist between the histological characteristics of our HCC cases and that of other populations.