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Optimized Purification of CIDRα-PfEMP1 Plasmodium falciparum Recombinant Protein with Affinity Chromatography Alami, Eqiel Navadz Akthar; Sulistyaningsih, Erma; Kusuma, Irawan Fajar; Rachmania, Sheilla
Jurnal ILMU DASAR Vol 23 No 2 (2022)
Publisher : Fakultas Matematika dan Ilmu Pengetahuan Alam Universitas Jember

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.19184/jid.v23i2.24127

Abstract

Interaction of Cysteine-rich Interdomain Region (CIDR)α-Plasmodium falciparum Erythrocyte Membrane Protein (PfEMP1) and Endothelial Cell Receptors especially CD36 on host cells is main malaria pathogenesis, makes this domain as a malaria vaccine candidate. Recently, the development of the malaria vaccine is conducted by recombinant technology, and the purification of the CIDRα-PfEMP1 recombinant protein is a pivotal step. This study aimed to determine an optimal condition to purify the CIDRα-PfEMP1 recombinant protein by affinity chromatography through imidazole and NaCl concentration. The purified recombinant protein was visualized using SDS-PAGE and its concentration was measured using Image J software and Bradford Assay. The data were analyzed using SPSS 26 software, and the Paired T-Test analysis was conducted to compare the concentration of purified recombinant protein from two different methods. The result showed that thetarget band of purified recombinant protein was 27 kDa. The thickest target protein band was observed in purified recombinant protein using 140 mM imidazole and 300 mM NaCl. The recombinant protein concentration using Image J software was 0.025 µg/µL, while the Bradford Assay was 0.56 µg/µL. The Paired T-Test analysis has a significance value of 0.010 (p<0.05), meaning there was a significant difference between the concentration measurement using Image J software and Bradford Assay. In conclusion, the optimized condition to purify the CIDRα-PfEMP1 recombinant protein by affinity chromatography was using 140 mM imidazole and 300 mM NaCl. It is suggested to measure the purified CIDRα-PfEMP1 recombinant protein concentration using the Bradford Assay method due to its convenience and sensitivity.
Analysis of Microglia Morphology and Number in Wistar Rats Brain After CIDR1α-PfEMP1 Recombinant Protein Injection Erma Sulistyaningsih; Izza Amalia Putri; Sheilla Rachmania
Al-Kauniyah: Jurnal Biologi Vol 18, No 1 (2025): AL-KAUNIYAH JURNAL BIOLOGI
Publisher : Department of Biology, Faculty of Science and Technology, Syarif Hidayatullah State Islami

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.15408/kauniyah.v1i1.38242

Abstract

AbstractOne malaria vaccine candidate is Cysteine-rich Interdomain Region 1α (CIDR1α) of Plasmodium falciparumErythrocyte Membrane Protein 1 (PfEMP1), an essential protein involved in the pathogenesis of cerebral malaria. Microglia in the brain act as the first line of defense against brain pathological changes. The study aimed to evaluate the response of brain microglia to the CIDR1α-PfEMP1 recombinant protein injection by observing microglia morphology and number in rat’s cerebral cortex. 12 Wistar rats were divided into the control group, which was injected with normal saline solution, and the treatment group, which was injected with 150 µg CIDR1α-PfEMP1 recombinant protein combined with adjuvants. Injection was conducted thrice within three-week intervals (day 1, 21, and 42). Wistar rats were euthanized on day 56, and histological slides were prepared with Hematoxylin-Eosin staining. Examination using a microscope, 400x, and Fiji Image J software showed microglia morphology of ramified and rod cells in both the control and treatment groups. The microglia number in the control group was 93.00 ± 5.77, and the treatment group was 105.75 ± 15.62. Statistical analysis using an independent t-test showed no significant differences between groups (p= 0.15). The result indicated that the injection of CIDR1α-PfEMP1 recombinant protein did not provoke pathological changes in brain tissue, which induced a microglia response. This study strengthens the potential of the CIDR1α-PfEMP1 recombinant protein as a peptide-based malaria vaccine candidate.AbstrakSalah satu kandidat vaksin malaria adalah Cysteine-rich Interdomain Region 1α (CIDR1α) dari Plasmodium falciparum Erythrocyte Membrane Protein 1 (PfEMP1), protein penting dalam patogenesis malaria serebral. Mikroglia di otak berperan sebagai pertahanan lini pertama terhadap perubahan di otak. Penelitian ini bertujuan mengevaluasi respon mikroglia otak terhadap pemberian protein rekombinan CIDR1α-PfEMP1 dengan mengamati morfologi dan jumlah mikrolia pada korteks serebri otak tikus. 12 tikus Wistar dibagi dalam kelompok kontrol yang diinjeksi normal saline dan  kelompok perlakuan diinjeksi 150 µg protein rekombinan CIDR1α-PfEMP1 yang dikombinasikan dengan adjuvant. Injeksi dilakukan tiga kali dengan interval tiga minggu (hari 1, 21, dan 42). Tikus dieuthanasia pada hari ke-56 dan preparat histologi otak disiapkan dengan pengecatan Hematoxyline-Eosin. Pengamatan menggunakan mikroskop 400x dan Fiji Image J software menunjukkan morfologi ramified dan rod cell pada kelompok kontrol maupun perlakuan. Jumlah mikroglia pada kelompok kontrol 93,00 ± 5,7 sedangkan kelompok perlakuan 105,75 ± 15,62). Analisis statistik menggunakan independent-t test menunjukkan tidak terdapat perbedaan yang bermakna antara 2 kelompok (p= 0,15). Hasil ini mengindikasikan bahwa pemberian protein rekombinan CIDR1α-PfEMP1 tidak menimbulkan patologi pada jaringan otak yang memicu respon mikroglia. Hal ini menguatkan potensi protein rekombinan CIDR1α-PfEMP1 sebagai kandidat vaksin malaria berbasis peptida.
Shallot Peel Infusion Prevents Bronchus Epithelium Thickening And Cilia Shortening In Cigarette Smoke-Induced Wistar Rats Bintang, Muhammad Farrel Ravidinata Masoga; Dewi, Rosita; Efendi, Erfan; Helianti, Dina; Rachmania, Sheilla
Jurnal Profesi Medika : Jurnal Kedokteran dan Kesehatan Vol 18 No 2 (2024): Jurnal Profesi Medika : Jurnal Kedokteran dan Kesehatan
Publisher : Fakultas Kedokteran UPN Veteran Jakarta Kerja Sama KNPT

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.33533/jpm.v18i2.7804

Abstract

Cigarette smoke exposure is the major risk of chronic obstructive pulmonary disease, the third cause of death worldwide. Inhaled smoke triggers oxidative stress resulting in airway epithelium thickening and cilia shortening. Shallot (Allium cepa L.) peel contains flavonoid which can neutralize oxidative stress. This study aims to determine correlation between shallot peel infusion (SPI), bronchus epithelium thickness, and cilia length in cigarette smoke-induced rats; and establish maximum effective dose of SPI. Rats were divided into normal; cigarette; SPI 125, 250, 500, 1,000, 2,000 mg/kgBW groups. Two hours after administration, rats were exposed to cigarette smoke 2 cigarettes/day for 28 days. Hematoxylin-eosin-stained bronchus was observed and variable measurements were carried out. Comparation tests of epithelium thickness and cilia length between normal and cigarette group showed significant difference (p<0.05); Pearson coefficient between SPI dose and epithelium thickening was -0.614, Spearman coefficient between SPI dose and cilia length was 0.860; and maximum effective dose to prevent bronchus epithelium thickening and cilia shortening are 1,275.4 mg/kgBW and 1,325.8 mg/kgBW. In conclusion, the higher the SPI dose, the lower epithelium thickness, the higher cilia length. Maximum effective dose of SPI to prevent bronchus epithelium thickening and cilia shortening are 1,275.4 mg/kgBW and 1,325.8 mg/kgBW.
EVALUATION OF BRAIN MICROGLIA PROLIFERATION AS A RESPONSE TO DBL2β-PFEMP1 RECOMBINANT PROTEIN IMMUNIZATION IN WISTAR RAT Dewi, Rosita; Kusuma, Irawan Fajar; Sulistyaningsih , Erma; Rachmania, Sheilla; Luthfiyyah, Nur Atikah
MNJ (Malang Neurology Journal) Vol. 11 No. 1 (2025): January
Publisher : PERDOSSI (Perhimpunan Dokter Spesialis Saraf Indonesia Cabang Malang) - Indonesian Neurological Association Branch of Malang cooperated with Neurology Residency Program, Faculty of Medicine Brawijaya University, Malang, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21776/ub.mnj.2025.011.01.04

Abstract

Background: In the malaria vaccine study, Duffy binding-like 2β Plasmodium falciparum erythrocyte membrane protein 1 (DBL2β-PfEMP1) could induce the  IgG and CD4+production. Antibody to DBL2β-PfEMP1 reduces the risk of developing severe malaria through the blockade of cytoadherence and destruction of rosette formation. During the process of antibody formation after immunization, the released peripheral cytokines have the potential to cause blood-brain barrier disruption resulting in the activation and proliferation of brain microglia as primary innate immune cells leading to neuroinflammation. Objective: This study aims to evaluate brain microglia proliferation as a response to recombinant protein DBL2β-PfEMP1 immunization in Wistar rats. Methods: Wistar rats were injected subcutaneously with recombinant protein DBL2β-PfEMP1 at doses of 100, 150, and 200 µg/kgBW on days 0, 21, and 42. Rats were euthanized on day 56. Brain histopathological slides were prepared and stained using hematoxylin-eosin. Histological examination was performed using a microscope at 400X magnification and documented using an AmScope microscope digital camera. Brain microglia were calculated using Fiji Image-J. The data were statistically analyzed using the Kruskal-Wallis test.  Results: The average number of brain microglia in both the control and treatment groups was 82–88. There was no significant difference in brain microglia number between the control and treatment groups (p>0.05) after recombinant protein DBL2β-PfEMP1 immunization. Conclusion: Recombinant protein DBL2β-PfEMP1 immunization does not provoke the proliferation of brain microglia in Wistar rats. This suggests that the protein does not have the potential to cause neuroinflammation.
In silico Investigation on Clopidogrel, Prasugrel and Ticagrelor as Potential Mono Antiplatelet Therapy for Acute Coronary Syndrome Hibatullah, Muhammad Naufal; Suryono, Suryono; Rachmania, Sheilla
Molecular and Cellular Biomedical Sciences Vol 8, No 3 (2024)
Publisher : Cell and BioPharmaceutical Institute

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21705/mcbs.v8i3.464

Abstract

Background: In acute coronary syndrome (ACS), antiplatelet therapy is crucial for inhibiting platelet aggregation. Dual antiplatelet therapy (DAPT) commonly employs aspirin along with clopidogrel, prasugrel, or ticagrelor. This is well known that aspirin acts as a cyclooxygenase (COX)-1 inhibitor, while clopidogrel, prasugrel, and ticagrelor act as P2Y12 inhibitors. Despite DAPT's proven efficacy in more effectively reducing cardiovascular events in ACS patients, this is associated with an increased risk of bleeding compared to mono antiplatelet therapy (MAPT). To minimize the cost and side effect that might arise from the use of DAPT, this is necessary to assess the potential of MAPT using a P2Y12 inhibitor drug, to understand whether they are capable of binding to both COX-1 and P2Y12. Hence, this study was conducted to identify P2Y12 inhibitor drugs that have the ability to bind to COX-1, allowing them to be proposed as MAPT.Materials and methods: Molecular docking was employed to assess binding affinity, interaction types, amino acid residues, binding distances, and visualizations in both 3D and 2D formats. The applications utilized were BIOVIA Discovery Studio and AutoDock, while the websites utilized were research collaboratory for structural bioinformatics protein data bank (RCSB PDB) and PubChem.Results: In silico findings reveal differences in binding strength among clopidogrel, prasugrel, and ticagrelor to COX-1 and P2Y12, with ticagrelor emerging as the stronger ligand due to a higher number of bindings and/or closer binding distances. Notably, only prasugrel and ticagrelor demonstrate the ability to bind to the active site of COX-1.Conclusion: Therefore, prasugrel and ticagrelor emerge as potential MAPT agents for ACS patients.Keywords: clopidogrel, prasugrel, ticagrelor, antiplatelet, ACS, in silico, molecular docking
Evaluation of Kidney Proximal Tubule Following Immunization with Plasmodium falciparum CIDR1α-PfEMP1 Recombinant Protein in Rats Dewi, Rosita; Sulistyaningsih, Erma; Kusuma, Irawan Fajar; Rachmania, Sheilla; Rahma, Nafisah Hani Asyifah
Majalah Kedokteran Bandung Vol 57, No 1 (2025)
Publisher : Faculty of Medicine, Universitas Padjadjaran

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.15395/mkb.v57.3801

Abstract

Malaria vaccines are continuously explored as an approach to eradicate malaria. The cysteine-rich interdomain region 1α-Plasmodium falciparum erythrocyte membrane protein 1 (CIDR1α-PfEMP1) is an antigenic protein that can bind to the endothelial protein C receptor (EPCR) and CD36, resulting in microvascular obstruction. The PfEMP1-induced antibody can induce antibodies, reducing the severity of malaria risk by impeding cytoadherence and destructing rosette formation. Preclinical safety testing is an important step of vaccine development, including safety testing of the kidney as the main excretory organ. The proximal tubule has the most mitochondria to support its main role in reabsorption and excretion, making it prone to oxidative stress caused by foreign substances. This study aimed to evaluate kidney proximal tubule cells after CIDR1α-PfEMP1 immunization in rats. This study was conducted at the Laboratory of Biology Molecular and Biotechnology, Faculty of Medicine, University of Jember. Eight rats were injected subcutaneously with 150 µg of the protein and four rats were injected with 0.9%  NaCl on days 0, 21, and 42. The rats were euthanized on day 56. The kidney histopathological slides were stained using Hematoxylin-Eosin (HE) and the necrotic proximal tubule cells were counted at five (5) visual fields (100 cells/visual fields). The average number of necrotic cells of the control and the treatment groups were 0.125±0.25 and 2.438±2.5972 while the Mann-Whitney test showed a significance value of p=0.12,  indicating no significant difference between the control and treatment groups. In conclusion, there is no change in the kidney histopathology based on the proximal tubule necrotic cell count after CIDR1α-PfEMP1 immunization in rats.
Differential Leukocyte Count Responses Post Injection of Duffy-binding-like Domain-2β of PfEMP1 Recombinant Protein in Wistar Rat Zahniar, Zahniar; Sulistyaningsih, Erma; Rachmania, Sheilla; Dewi, Rosita; Kusuma, Irawan Fajar
Medical Laboratory Technology Journal Vol. 11 No. 1 (2025): June
Publisher : Poltekkes Kemenkes Banjarmasin Jurusan Analis Kesehatan

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.31964/mltj.v11i1.628

Abstract

Malaria due to Plasmodium falciparum causes a high mortality rate, and vaccination is a valuable approach to control it. One malaria vaccine candidate is Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP1), specifically Duffy binding-like 2β (DBL2β) domain (DBL2β-PfEMP1), which has a vital role in severe malaria pathogenesis. The DBL2β-PfEMP1 recombinant protein is immunogenic. This study aimed to investigate the immune response of DBL2β-PfEMP1 protein by analyzing the differential leukocyte count. Twenty-three rats were randomly divided into control and five treatment groups. Rats were injected on days 0, 21, and 42 with a physiological solution of 0.9% NaCl, adjuvant, DBL2β-PfEMP1 protein, and each mixture of DBL2β-PfEMP1 protein with doses of 150, 300, and 450 µg/200gBW and adjuvant. Blood was collected on day 56 and prepared for differential leukocyte count examination with a visual microscopic examination by two expert observers. The results showed that DBL2β-PfEMP1 recombinant protein and adjuvant increased the eosinophils and neutrophils but decreased monocytes and lymphocytes and did not affect the basophils. Statistical analysis showed significant differences between groups for eosinophils (between control and DBL groups; Adj and DBL groups; DBL and other groups except DBL150+adj) and monocytes (between control and all doşe groups with adjuvant; DBL and all doşe groups with aduvant), but not for basophils, neutrophils, and lymphocytes. In conclusion, the serial injection of DBL2β-PfEMP1 recombinant protein showed different responses in each leukocyte cell type. Further analysis by time-series differential leukocyte count examination will be essential to determine the responses of each type of leukocyte to support the research on malaria vaccine development.
Relationship between PLR Value and Severity of Dengue Infection in Pediatric Patients in the Aster Room at RSUD dr. Soebandi Jember Gianevan, Nicholas Jirezra; Riyanti, Rini; Shodikin, Muhammad Ali; Rachmania, Sheilla; Dewi, Rosita
Journal of Agromedicine and Medical Sciences Vol. 11 No. 2 (2025)
Publisher : Faculty of Medicine, Universitas Jember

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.19184/ams.v11i2.50026

Abstract

Dengue infection is a disease caused by dengue virus infection and transmitted by the Aedes mosquito, and is a major health problem in children, especially in developing countries. The Platelet to Lymphocyte Ratio (PLR) is a novel inflammatory biomarker that has recently been frequently studied and plays a role in the assessment of viral infectious diseases, and the use of PLR in viral infectious diseases has previously found greater potential compared with the Neutrophil-Lymphocyte Ratio (NLR). It is hoped that PLR can become a new biomarker for predicting the severity of dengue infection which is suitable for use in developing countries because PLR ​​is a cheap biomarker and is available in all clinical conditions. This research is a correlative analytical observational study using a cross-sectional approach. This research design was carried out by collecting data from children diagnosed with dengue infection as well as data on platelets and lymphocytes on the first day of admission to the hospital. This research uses stratified random sampling. Data were analyzed using Spearman correlation with a confidence level of 99%. The results of Spearman's correlation analysis showed a strong significant relationship with p<0.01 (p=0.000) and a correlation coefficient of -0.753. In conclusion, there is a significant and strong relationship between the Platelet-to-Lymphocy Ratio (PLR) value and the severity of dengue infection in pediatric patients in the Aster ward of RSUD Dr. Soebandi Jember. If the PLR ​​value increases, the severity of dengue infection will be lower or decrease and vice versa. Keywords: PLR; dengue infection; children; platelets; lymphocytes
Tobacco Dust Exposure Among Tobacco Workers in Jember Reduces Lung Function Capacity Nofica, Cheila; Firdaus, Jauhar; Rachmania, Sheilla; Caesarina, Ancah
International Journal of Health and Information System Vol. 3 No. 2 (2025): September
Publisher : Indonesian Journal Publisher

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.47134/ijhis.v3i2.60

Abstract

This study investigated the association between urinary cotinine levels and lung function among tobacco workers in Jember Regency, a major tobacco-producing region in Indonesia. A total of 48 respondents were included based on specific inclusion and exclusion criteria. Urinary cotinine levels were measured using enzyme-linked immunosorbent assay (ELISA), and lung function was assessed via spirometry. Based on FEV1/FVC ratios, 58% of participants exhibited restrictive lung patterns, 29% showed obstructive patterns, and 13% had normal pulmonary function. Statistical analysis conducted using SPSS, with a significance level set at p < 0.05, indicated no statistically significant correlation between urinary cotinine levels and lung function.
Molecular Method Optimization to Identify Plasmodium falciparum Multidrug Resistance 1 (pfmdr1) gene as a Predictor of Antimalarial Resistance Sulistyaningsih, Erma; Dewi, Rosita; Rachmania, Sheilla; Kusuma, Irawan Fajar; Kholifaturrohmy, Muhammad Rizqi; Armiyanti, Yunita; Kholifaturrohmah, Sakinatus Sariroh; Andriani, Made Prasanti
Biology, Medicine, & Natural Product Chemistry Vol 14, No 2 (2025)
Publisher : Sunan Kalijaga State Islamic University & Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14421/biomedich.2025.142.673-678

Abstract

Several approaches have been designed to control malaria, a disease with high morbidity and mortality, but they face some hurdles. Antimalarial resistance is one of the major challenges for malaria elimination, so the detection of antimalarial resistance is essential. Several molecular markers for antimalarial resistance have been identified, including Plasmodium falciparum multidrug resistance 1 (pfmdr1) gene. This study determined the optimization of molecular techniques to identify the pfmdr1 gene as an antimalarial resistance predictor in Indonesia. The study included patients diagnosed with uncomplicated or severe malaria originating from the health district of Kerom Regency, Papua Province, and Dr. Soebandi Hospital, Jember, East Java Province. Blood samples were collected in the Whatmann filer paper after informed consent. DNA was isolated from dried blood filter paper, and nested PCR was performed using a specific primer, the pfmdr1-A and pfmdr1-B genes. The PCR cycle was optimized based on previous studies. The pfmdr1-A has a similar setting to the earlier study, but the pfmdr1-B had a different optimum setting from the previous study with the annealing temperature of 57oC for nested-1 and 62oC for nested-2. This PCR setting could be used for further examination. The positive results of the amplification indicated the potential for antimalarial resistance in the parasite population. A study on the gene copy number and polymorphism is essential to determine the definitive conclusion on antimalarial resistance.