The Indonesian Biomedical Journal

242-55

Authors: Chandra Agung Purnama, Anna Meiliana, Melisa Intan Barliana, Keri Lestari Dandan, Andi Wijaya

BACKGROUND: Millions of cells in multicellular organisms regenerate every day to replace aged and died cells. Effective cell clearance (efferocytosis) is critical for tissue homeostasis, as the human body recycles its cellular components. We summarize what is known about the mechanisms of efferocytosis and how it impacts the physiology of the organism, effects on inflammation and the adaptive immune response, as well as the consequences of defects in this critical homeostatic mechanism in this review. CONTENT: Cell death is the process by which the human body replaces aged or damaged cells with new ones. It can be triggered by genetically encoded machinery or regulated cell death, or by specific pharmacologic or genetic interventions, resulting in accidental cell death. Dying cells release signals that entice phagocytes to engulf them in a process known as efferocytosis. Efferocytosis is a multistep process involving the release of “find me” and “eat me” signals and destruction of death cells by phagocytes. Different types of cell death including apoptosis and necroptosis can express pro- or anti-inflammatory signals via macrophage activity modulation. SUMMARY: Failed or ineffective efferocytosis can result in disruption of tissue homeostasis, which can contribute to the development of chronic inflammatory diseases such as atherosclerosis, obesity, diabetes, and heart failure. Therefore, any therapeutic strategy that enhances efferocytosis will have a beneficial effect on the treatment of these metabolic disorders.

261-70

Authors: Aigerim Turgimbayeva, Sailau Abeldenov, Nurgul Sarina, Bekbolat Khassenov, Saule Eskendirova

BACKGROUND: Active proliferation is specific property of a tumor cells. However, the cost of the analysis is high due to commercial anti-Ki-67 mAbs used as the main immunoreagent for reliable identification of proliferating cells. In this study, recombinant protein was used to obtain specific mAbs for Ki-67 biomarker immunodetection. METHODS: Codon optimized fragment of ki-67 gene was cloned into the pET28c(+)vector. The recombinant protein was purified by immobilized metal affinity chromatography (IMAC) and confirmed by liquid chromatography–mass spectrometry (LC-MS)/MS. Hybridoma cells were obtained by fusing myeloma cells with mouse spleen cells immunized with recombinant antigen. The specificity and activity of mAbs was determined by enzyme-linked immunosorbent assay (ELISA), Western blot and immunocytochemistry. RESULTS: The pET-28c(+)/ki-67 plasmid, which encodes 355 amino acid protein, was obtained. Analysis by LC-MS/MS of the recombinant antigen showed that 77.5% of the amino-acid sequence belonged to Ki-67 protein. Recombinant fragment of Ki-67 protein was used to obtain specific hybridoma strains. ELISA and Western blot demonstrated high affinity and the specificity of obtained mAbs against Ki-67 protein. Newly generated anti-Ki67 mAbs detected target protein in proliferating cells of MCF-7 cell line by immunocytochemistry. CONCLUSION: Newly developed mAbs are potentially useful as an immunodiagnostic tool for assessing the proliferative activity of breast tumor cells using immunocytochemistry.

281-8

Authors: Dewinda Candrarukmi, Annang Giri Moelyo, Muhammad Riza

BACKGROUND: Hyperglycemia is one of the most common endocrine complications in children with β thalassemia major. Though the current diagnostic marker either requires fasting, has low reproducibility, or it is not an accurate for thalassemia patients. Glycated albumin (GA) is a quick and easy alternative marker for hyperglycemia detection and monitoring of glycemic control. However to date, no studies have analyzed the role of GA value in detection of hyperglycemia in children with thalassemia major. This study analyzed the value of GA as an alternative screening marker for hyperglycemia detection in children with β thalassemia major. METHODS: A single-blind prospective diagnostic test was conducted in 9 to 18 years old children with β thalassemia major and who were treated at the Dr. Moewardi Regional General Hospital between October 1, 2018 and December 31, 2019. In a single, fasted study visit, height, weight, fasting blood glucose (FPG), GA, oral glucose tolerance test (OGTT) were measured. The area under a receiver operating characteristic curve (AUC) was used to determine the cut-off value at which hyperglycemia prediction (OGTT ≥140 mg/dL) display optimal sensitivity and specificity. RESULTS: Among the 53 children with β thalassemia major, 1 (1.9%) had diabetes mellitus and 4 (7.5%) had prediabetes based on their OGTT values. The median GA value in this study was 10.9% (range: 7.6–12.4%). GA had a low AUC (0.646, p=0.287) for hyperglycemia detection in pediatric patients with β thalassemia major. At a cut-off of 11.45%, GA demonstrated 60% sensitivity and 60.4% specificity. CONCLUSION: GA cannot be used as an alternative marker for hyperglycemia detection in children with β thalassemia major.

295-302

Authors: Dwita Permatasari, Hendrian Dwikoloso Soebagjo, Ismi Zuhria, Nila Kurniasari, Hari Basuki Notobroto, Cinintha Nandini

BACKGROUND: Usually, surgical intervention is needed to eradicate the fungal microorganism that cause fungal corneal ulcers. However, since surgical intervention is invasive, the latest technology uses cryotherapy in treating it. Cryotherapy plays a vital role in the wound healing process. We aimed to evaluate is to study the decreased expression of matrix metalloproteinase 9 (MMP-9) and transforming growth factor β1 (TGF-β1) in fungal corneal ulcers after the administration of cryotherapy. METHODS: Aspergillus flavus fungus was injected to the intrastromal corneas of all Sprague Dawley rats. The rats were divided into four groups, the first group was not given any therapy, the second group was given topical natamycin therapy, the third group was given cryotherapy, and the fourth group was given a combination between cryotherapy and topical natamycin therapy. Therapy was given after five days of follow up on the formation of a corneal ulcer. After four days of therapy, the eyes were enucleated to determine MMP-9 and TGF-β1 expression. RESULTS: The result in the third group showed lower MMP-9 expression (20.0±10.0% cells per field of view) compared to the second group (40.0±20.0% cells per field of view) and the fourth group (30.0±25.0% cells per field of view), but had the same MMP-9 expression value as the first group. There was no significant difference in MMP-9 expression between the four groups (p=0.356). The third group reduced more TGF-β1 expression (10.0±12.50% cells per field of view) compared to the fourth group (30±27.5% cells per field of view) and the first group (30±32.5% cells per field of view). There was also no significant difference in TGF-β1 between the four groups (p=0.315). CONCLUSION: There is no significant difference in the expression of TGF-β1 and MMP-9 after the cryotherapy treatment.

310-5

Authors: Fatmawati Fatmawati, Ellyza Nasrul, Nasrul Zubir, Ferry Sandra

BACKGROUND: T lymphocyte activation depends on the balance of co-stimulatory and co-inhibitory signals determined by Cluster of Diffrentiation (CD)28 and Programmed Cell Death Protein 1 (PD-1) expression. Alteration in CD28 and PD-1 expression might affect the progression of chronic hepatitis B (CHB). Current study was conducted to evaluate the correlations of the CD28 and PD-1 expressions of T lymphocytes and CHB progression. METHODS: Subjects were recruited, selected and divided into 3 groups, inactive CHB, active CHB and CHB with End-Stage Liver Disease (ESLD). HBeAg was determined by using Enzyme-Linked Fluorescence Assay while HBV-DNA was carried out by the RT-PCR method. Numbers of T lymphocytes expressing CD3, CD4, CD8, CD45, CD28 and PD-1 molecules were determined by flowcytometry. RESULTS: There was no significant difference in the expression of CD28 by CD4+ and CD8+ T lymphocytes of inactive CHB, active CHB and CHB with ESLD subjects. There was also no significant difference in the expression of PD-1 in CD4+ lymphocytes of inactive CHB, active CHB and ESLD subjects. In contrast there was a significant increase in the expression of PD-1 in CD8+ T lymphocytes of ESLD subjects. CONCLUSION: CD28 expression among CHB subjects was within normal range and not related to disease progression, but PD-1 expression of CD8+ T lymphocyte was increased along with disease progression, especially in CHB subjects with ESLD. This suggests that PD-1-overexpressed CD8+ T lymphocyte play a role in increasing CHB disease progression.

324-31

Authors: Thianti Sylviningrum, Ismiralda Oke Putranti, Octavia Permata Sari, Fitranto Arjadi, Triasari Oktavriana

BACKGROUND: Psoriasis is an autoimmune disease involving genetic-environmental factors. Human Leukocyte Antigen (HLA)-Cw6 allele is the main genetic risk factor of psoriasis, but its prevalence varies widely. HLA-Cw6 allele correlates with psoriasis clinical type and treatment responses. Narrowband ultraviolet B (NB-UVB) and methotrexate (MTX) are effective psoriasis treatments but their association with HLA-Cw6 allele has not been identified. The study aims to determine the association between HLA-Cw6 allele expression, NB-UVB, and MTX treatment responses in Javanese-Indonesian psoriasis subjects. METHODS: Ninety Javanese-Indonesian psoriasis subjects were recruited in this study, 45 subjects were treated using NB UVB, while the other 45 subjects were treated using MTX, respectively, for 12 weeks. The psoriasis diagnosis and treatment responses were evaluated by dermatologists using Psoriasis Area Severity Index (PASI) score. The HLA-Cw6 allele was examined using the single-specific-primer polymerase chain reaction method. Fisher's Exact and Chi-square tests were employed, where p-value<0.05 was considered of significant association. RESULTS: The HLA-Cw6 allele positivity was identified in 23 psoriasis subjects (25.56%), while the other 67 subjects expressed HLA-Cw6 allele negative (74.44%). Female with HLA-Cw6 allele positivity who did not have comorbid disease show good response to NB-UVB than MTX. Meanwhile, subjects who were treated with MTX showed no association between therapeutic response and HLA-Cw6. HLA-Cw6 status was not correlated with the onset of psoriasis, family history, and comorbid diseases in all subjects. CONCLUSION: HLA-Cw6 allele expression is more associated with good NB-UVB treatment response than with MTX treatment response in female Javanese-Indonesian psoriasis subjects without comorbid disease.