A Modified Method To Create A Porcine Deep Dermal Burn Model.

Development of an experimental burn model is essential to studying burn wound pathophysiology and progressivity, and identifying the best therapeutic choice before treatment can be applied clinically. Pigs are preferred as animal models for burns because their skin structure and wound regeneration resemble that of humans. We made a novel model to create a deep dermal burn using prospective methods that are easy to use, safe, and consistently reproducible. A custom-made burn device was used to create deep dermal burn wounds. A cylindrical plate made of stainless steel with a diameter of 2cm was connected to an electric heating device that was set to reach temperatures varying between 82-100°C. The hot cylindrical plate was applied perpendicular to the pig's skin with a pressure of 1kgf mechanical force for various periods of time (10-25s). Burns were created on the dorsum of a male Yorkshire pig sedated under anesthesia and analgesia. At the 7th day post burn, the burn wounds were evaluated macroscopically, and when one fulfilled the criteria for deep dermal burn, tissue was harvested for further histological analysis using hematoxylineosin staining. At the end of the study, the burn wounds created by the temperature 92°C for 20s showed tissue damage that meets the criteria for deep dermal burns. The development of this model might be useful for further study on burn wound healing.

La mise au point de méthodes de brûlure expérimentale est essentielle pour l'étude de la physiopathologie et de l'évolution des brûlures, afin d'évaluer les meilleures options thérapeutique. La peau du porc a une structure et un processus de cicatrisation proches de la peau humaine, ce qui fait de sa brûlure un modèle particulièrement fiable. Nous avons développé un modèle de brûlure porcine du 2ème degré simple, sécuritaire et aisément reproductible. Un cylindre d'acier inoxydable de 2 cm de diamètre et connecté à un chauffage électrique réglable entre 82 et 100°C. Il est appliqué perpendiculairement à la peau, sous une pression de 1 kg pendant une durée réglable de 10 à 25 s. Les modèles sont des porcs Yorkshire mâles, la brûlure est infligée sous AG. À J7, les brûlures sont examinées au microscope. Lorsque l'examen confirme l'atteinte au 2ème degré profond, le tissu est conservé pour examen ultérieur après coloration hématoxylineéosine, qui confirme que 20s à 92°C sont nécessaires pour créer ce type de lésion. Ce modèle pourrait servir de base aux futures études sur la cicatrisation des brûlures.

Comparison of multiplex ligation dependent probe amplification to immunohistochemistry for assessing HER-2/neu amplification in invasive breast cancer.

The HER-2/neu transmembrane tyrosine kinase receptor is both a prognostic marker and a therapeutic target for breast cancer. Accurate determination of HER-2/neu status is a prerequisite for selecting breast tumors for HER-2/neu immunotherapy or for taxan based chemotherapy. Unfortunately, there is no consensus concerning how this determination should be reached. We compared assessment of HER-2/neu status using Multiplex ligation-dependent probe amplification (MLPA) and immunohistochemistry (IHC). The patient group comprised 60 Indonesian breast cancers patients. IHC was performed on paraffin sections using the CB11 antibody from Novocastra. Results were scored according to the Hercept test. For MLPA, DNA was extracted from frozen samples, PCR amplified with a probe set containing three hemi-primer sets for the HER-2 locus and another nine control probes spread over chromosome 17 and other chromosomes, and analyzed on a gene scanner. A ratio above two for at least two HER-2 locus probes compared to the control probes was regarded as amplification. IHC for HER-2/neu was negative in 36 cases, and 24 cases (40%) showed expression. Seven, eight and nine of the latter cases were 1+, 2+ and 3+ positive, respectively. Forty-seven cases showed no amplification by MLPA, and 13 cases (22%) were amplified. Comparison of IHC and MPLA showed that none of the 36 IHC-negative or seven IHC 1+ cases was amplified. Five of the eight (63%) 2+ cases were amplified, and eight of nine (89%) of the IHC 3+ tumors showed gene amplification by MLPA assay. For HER-2/neu, there is a good correlation between gene amplification detected by MLPA and overexpression by IHC in invasive breast cancer. It appears that MLPA can detect the HER-2 amplified cases in the IHC 2+ class. Because MLPA is quick and inexpensive, it is an attractive method for detecting HER-2/neu amplification in daily laboratory practice.

A novel BRCA2 mutation in an Indonesian family found with a new, rapid, and sensitive mutation detection method based on pooled denaturing gradient gel electrophoresis and targeted sequencing.

BACKGROUND: Breast cancer is increasing in Indonesia and other developing countries. Germline mutations in the BRCA1/2 genes are most strongly associated with a high risk for breast cancer development. There have been no reports on BRCA1/2 gene mutations in the Indonesian population. Genetic research yielding insight into mutations affecting the Indonesian population can help in risk assessment of individual patients. AIMS: To screen the BRCA1/2 genes for mutations in early onset Indonesian breast cancer patients and their families with a new, simple, and sensitive BRCA1/2 mutation screening strategy based on denaturing gradient gel electrophoresis (DGGE) and targeted sequencing. METHODS: DNA was isolated from the blood of four Indonesian breast cancer patients from high risk families and seven family members, and the polymerase chain reaction was performed with specially designed primers throughout the BRCA1/2 coding sequences to produce fragments suitable for pooled DGGE analysis. The aberrantly migrating samples were reamplified and sequenced. RESULTS: Two mutations were found in exons 13 and 16 of BRCA1 and two mutations in exons 2 and 14 of BRCA2, which turned out to be established polymorphisms according to the Breast Cancer Information Core. In addition, a novel 6 bp deletion in exon 11, leading to a premature stop, was found in BRCA2. CONCLUSION: Pooled DGGE and targeted sequencing revealed four BRCA1/2 polymorphisms and one novel BRCA2 mutation in a group of Indonesian patients at high risk of hereditary breast cancer. This illustrates that the proposed method is sensitive and particularly suited for screening unknown populations.

Correlation of clinical, pathological status, hormone receptor and C-erbB-2 oncoprotein in breast cancer patients.

This study is to evaluate the correlation of some established prognostic factors, hormone receptor and C-erbB-2 expression of breast cancer patients in Yogyakarta, Indonesia. Beginning January 1997, 60 breast cancer patients who were treated either by mastectomy, breast conserving surgery or biopsy, were evaluated clinically in connection with age, menopausal status, stage, tumor size, nodes; also histologically regarding type, grade and mitotic index. Patients were evaluated for estrogen and progesterone receptor, as well as C-erbB-2 expression with immunohistochemical techniques. Median age was 47.5 years old, range from 28 to 80 years old. Most of them were premenopause (65.0%). One patient (1.7%) was a man. Most of the patients were stage IIB (25.0%), 51.7% with positive estrogen receptor and 65.9% with positive progesterone receptor. The type was mostly invasive duct carcinoma, high grade (70.0%). Most of the tumor size ranged between 2-5 cm (56.9%), with more than 3 nodes in 38.3% of patients. High mitotic index was found in 69.5% and positive C-erbB-2 in 71.7% patients. Correlation of C-erbB-2 and other prognostic factors showed that only stage, node and mitotic index had significant correlations (p = 0.016; 0.035 and 0.005, respectively). A significant correlation was also found between ER and PR level, PR and tumor size, stage and tumor size, stage and nodes, tumor size and nodes (p < 0.05), and a borderline correlation between ER and tumor size (p = 0.065) in conclusion, this preliminary study showed that breast cancer in Yogyakarta had an aggressive phenotype. C-erbB-2 positivity was correlated significantly with stage of the disease, number of nodes involved and mitotic index. Hormone receptors also correlated with some prognostic factors in breast cancer patients.