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Detection of the human papillomavirus in cervical carcinoma: a comparison between non-isotopic in-situ hybridisation and polymerase chain reaction as methods for detection in formalin-fixed, paraffin-embedded tissues

Cheah PL, Looi LM

Malays J Pathol, 2008 Jun;30(1):37-42.
PMID: 19108410

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.

Matched MeSH terms: Paraffin Embedding
Immunostaining of formalin-fixed, paraffin-embedded tissues for malignant lymphomas

Lim YC, Phang KS, Cheong SK

Malays J Pathol, 1992 Dec;14(2):85-9.
PMID: 1304629

With the advent of new monoclonal antibodies that are applicable to formalin-fixed, paraffin embedded sections, immunophenotyping is becoming increasingly important in the diagnosis and classification of lymphomas. However, multiple factors such as fixation, trypsinization and even type of antibodies used have certain effects on the final outcome of the staining procedure. In this paper we report our experience and the problems encountered in our laboratory when we first tried to establish a workable immunostaining protocol for formalin-fixed, paraffin embedded tissue sections using the immunoalkaline phosphatase technique.

Matched MeSH terms: Paraffin Embedding*
Expression of IGFBP-rP1 in ovarian and breast cancers in association with diabetes mellitus status

Mohd Nafi SN, Siti Azrin AH, Mat Zin AA, Othman NH, Che Jalil NA

Malays J Pathol, 2019 Apr;41(1):33-39.
PMID: 31025635

INTRODUCTION: Insulin-like growth factor binding protein-related protein 1 (IGFBP-rP1) is an important component of the IGF system that regulates insulin resistance-related to tumour development. The aim of this study is to investigate the expression of IGFBP-rP1 among female cancer patients who are known or not known to have Type 2 Diabetes Mellitus (T2DM).
MATERIALS AND METHODS: Using a cross-sectional design, cases of ovarian and breast cancer with clinical status of T2DM were selected over a 10-year period in Hospital Universiti Sains Malaysia. Immunohistochemical staining for IGFBP-rP1 was performed on paraffin-embedded tissues and the results were correlated with the patient's demographic and clinicopathological data.
RESULTS: A total of 152 breast cancer patients were recruited into the current study with 33.5% (51/152) patients were positive T2DM. Most of the breast cancer patients with T2DM were IGFBP-rP1-negative (66.7%, 34/51). The IGFBP-rP1 expression was significantly difference between breast cancer subjects with and without T2DM (p<0.001). There was no significant association of IGFBP-rP1 expression with data on the demographic and clinicopathological profiles of patients with breast cancer. Meanwhile, positive IGFBP-rP1 expression was evident in 44 out of 108 (40.74%) ovarian cancer cases. Among these cases, 36 were T2DM. In contrast to breast cancer cases, IGFBP-rP1 was mostly expressed among ovarian cancer patients with T2DM (66.7%, 24/36, p < 0.001). However, the -positive expression was not significantly associated with any sociodemographic and clinicopathological features of ovarian cancers.
CONCLUSIONS: Majority of breast cancer patients with T2DM did not express IGFBP-rP1. In contrast, majority of the ovarian cancer patients with T2DM expressed IGFBP-rP1.

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Fulltext Evaluation of DNA and RNA extraction methods

Edwin Shiaw CS, Shiran MS, Cheah YK, Tan GC, Sabariah AR

Med J Malaysia, 2010 Jun;65(2):133-7.
PMID: 23756798 MyJurnal

This study was done to evaluate various DNA and RNA extractions from archival FFPE tissues. A total of 30 FFPE blocks from the years of 2004 to 2006 were assessed with each modified and adapted method. Extraction protocols evaluated include the modified enzymatic extraction method (Method A), Chelex-100 extraction method (Method B), heat-induced retrieval in alkaline solution extraction method (Methods C and D) and one commercial FFPE DNA Extraction kit (Qiagen, Crawley, UK). For RNA extraction, 2 extraction protocols were evaluated including the enzymatic extraction method (Method 1), and Chelex-100 RNA extraction method (Method 2). Results show that the modified enzymatic extraction method (Method A) is an efficient DNA extraction protocol, while for RNA extraction, the enzymatic method (Method 1) and the Chelex-100 RNA extraction method (Method 2) are equally efficient RNA extraction protocols.

Matched MeSH terms: Paraffin Embedding*
Classifying DLBCL according cell of origin using Hans algorithm and its association with clinicopathological parameters: A single centre experience

Wn Najmiyah WAW, Azlan H, Faezahtul AH

Med J Malaysia, 2020 03;75(2):98-102.
PMID: 32281588

INTRODUCTION: In recent years, "double hit" and "double protein" involving gene rearrangement and protein expression of c-MYC and BCL2 and/or BCL6 are the most used terms to describe poor prognostic factors in diffuse large B-cell lymphoma (DLBCL). This study was to determine the frequency of double or triple protein expression by using immunohistochemistry (IHC) and comparing the result with clinicopathological features and cell of origin (COO) classification.
METHODS: We conducted a cross-sectional study by using 29 archived formalin-fixed paraffin embedded tissue blocks of DLBCL. All the samples were evaluated for the subgrouping of COO DLBCL was determined by expression of CD10, BCL6 and MUM1 based on Hans classification. In addition, expressions of c-MYC, BCL2 and BCL6 were detected by IHC.
RESULTS: Among the 29 cases, MYC, BCL2 and BCL6 proteins were detected in 72.4%, 62.1% and 62.1% of patients, respectively. Concurrent expression (c-MYC positive/BCL2 positive and/or BCL6 positive) was present in 58.6% of patients. 34.5% were categorised as germinal centre like (GCB) subgroup and 65.5% were categorised as nongerminal centre like (non-GCB) subgroup. Among the clinicopathological features, the double/triple protein expression lymphoma was significantly associated with elevated LDH level (p=0.018), IPI score (p=0.003), Ann Arbor stage (p=0.011) and complete response rate (p=0.011).
CONCLUSION: Double/triple protein lymphoma was strongly associated more adverse clinical risk factors. Thus, analyses of MYC, BCL2 and BCL6 expression by IHC represents a rapid and inexpensive approach to risk-stratify patients with DLBCL at diagnosis.

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Endocan expression in placenta of women with hypertension

Chew BS, Ghazali R, Othman H, Ismail NAM, Othman AS, Laim NMST, et al.

J Obstet Gynaecol Res, 2018 Oct 10.
PMID: 30306675 DOI: 10.1111/jog.13836

AIM: The aim of our study was to determine the endocan-1 expression in placenta of hypertensive women, and its association with maternal and fetal outcomes.
METHODS: This was a cross-sectional study consisted of 21 pregnant women with hypertension and 23 without hypertension. The gestational age ranged from 28 to 39 weeks (hypertensive) and 32 to 40 weeks (normotensive). The paraffin embedded formalin fixed placenta tissue blocks were retrieved from the pathology archives. Endocan immunohistochemistry was performed on tissue sections of full thickness and maternal surface of the placenta. The endocan expression was determined in fetal endothelial cells, maternal endothelial cells, cytotrophoblasts, syncytiotrophoblasts and decidual cells. The differences in endocan expression in placenta between hypertensive and normotensive subjects were evaluated by Pearson chi-square test and t-test were used in the statistical analysis.
RESULTS: The endocan expression was significantly higher in fetal endothelial cells (P

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Fulltext Neuronal Cell Death and Mouse (Mus musculus) Behaviour Induced by Bee Venom

Oktiansyah R, Juliandi B, Widayati KA, Juniantito V

Trop Life Sci Res, 2018 Jul;29(2):1-11.
PMID: 30112137 DOI: 10.21315/tlsr2018.29.2.1

Neuronal cell death can occur in a tissue or organ, including the brain, which affects memory. The objectives of this study were to determine the dose of bee venom that causes neuronal death and analyse the alteration of mouse behaviour, focusing in particular on spatial memory. Fifteen male mice of Deutsche Denken Yoken (DDY) strain were divided into control and treatment groups. Bee venom was injected six times for two weeks intraperitoneally with 1.88 mg/kg, 3.76 mg/kg, 5.6 mg/kg, and 7.48 mg/kg doses of venom. Brain histology was studied using haematoxylin-eosin stained paraffin embedded 5 μm coronal sections. A Y maze test was used to assay behaviour. Parameters observed were the number of dead neurons and the percentage of mice with altered behaviour. ANOVA showed that the effects of bee venom were significantly different in the case of the neuronal death parameter but were not significantly different in the case of the mice behaviour parameter. Duncan's Multiple Range Test (DMRT) demonstrated that P4 (7.48 mg/kg) gave the highest effect of bee venom to promote neuronal death.

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Fulltext A combined human case of Dirofilaria ursi infection in dorsal subcutaneous tissue and Anisakis simplex sensu stricto (s.s.) infection in ventral subcutaneous tissue

Yamada M, Shishito N, Nozawa Y, Uni S, Nishioka K, Nakaya T

Trop Med Health, 2017;45:26.
PMID: 29118653 DOI: 10.1186/s41182-017-0067-4

Background: Dirofilaria ursi is a filarial nematode that parasitizes the subcutaneous tissues of the American black bear (Ursus americanus) and Japanese black bear (Ursus thiabetanus japonicus). D. ursi that has parasitized black bears has the potential to subsequently infect humans. In addition, extra-gastrointestinal anisakiasis is less common in Japan.

Case presentation: We report a case of ventral subcutaneous anisakiasis and dorsal subcutaneous dirofilariasis that was acquired in Fukushima, in the northern part of Japan. The patient was an 83-year-old Japanese female, and subcutaneous parasitic granulomas were present on her left abdomen (near the navel) and left scapula. A pathological examination of the surgically dissected tissue sections from each region demonstrated eosinophilic granulomas containing different species of parasites. To enable the morphological and molecular identification of these parasites, DNA was extracted from paraffin-embedded sections using DEXPAT reagent, and the cytochrome oxidase 2 (COX2), internal transcribed spacer 1 (ITS1), 5.8S and ITS2 regions of the Anisakis larvae, and the 5S rRNA region of the male Dirofilaria were sequenced. The PCR products were examined and compared with DNA databases. Molecular analysis of the COX2 and 5S rRNA sequences of each worm revealed that the nematode found in the ventral region belonged to Anisakis simplex sensu stricto (s.s.) and the male Dirofilaria found in the dorsal region was classified as D. ursi.

Conclusion: The present case showed a combined human case of D. ursi and A. simplex s.s. infections in subcutaneous tissues. The results of this study will contribute to the identification of unknown parasites in histological sections.

Matched MeSH terms: Paraffin Embedding