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Fulltext Fatal case of amoebic liver abscess in a child

Ng KF, Tan KK, Ngui R, Lim YA, Amir A, Rajoo Y, et al.

Asian Pac J Trop Med, 2015 Oct;8(10):878-80.
PMID: 26522307 DOI: 10.1016/j.apjtm.2015.09.018

We reported a case of amoebic liver abscess (ALA) in a 6-year-old Malaysian boy who presented with fever, lethargy, diarrhoea and right hypochondriac pain. On admission he was diagnosed with perforated acute appendicitis and a laparotomy was done. After surgery he developed acute respiratory distress. Ultrasonography, chest X-Ray and CT scan revealed two ALAs in the posterior segment of right lobe of liver, pleural effusion and collapsed consolidation of lungs bilaterally. Percutaneous liver abscesses drainage was done and intravenous Metronidazole was started. PCR carried out on the pus from the abscess was positive for Entamoeba histolytica. Patient however succumbed to the infection one week after admission.
Fulltext Independent Emergence of the Cosmopolitan Asian Chikungunya Virus, Philippines 2012

Tan KK, Sy AK, Tandoc AO, Khoo JJ, Sulaiman S, Chang LY, et al.

Sci Rep, 2015 Jul 23;5:12279.
PMID: 26201250 DOI: 10.1038/srep12279

Outbreaks involving the Asian genotype Chikungunya virus (CHIKV) caused over one million infections in the Americas recently. The outbreak was preceded by a major nationwide outbreak in the Philippines. We examined the phylogenetic and phylogeographic relationships of representative CHIKV isolates obtained from the 2012 Philippines outbreak with other CHIKV isolates collected globally. Asian CHIKV isolated from the Philippines, China, Micronesia and Caribbean regions were found closely related, herein denoted as Cosmopolitan Asian CHIKV (CACV). Three adaptive amino acid substitutions in nsP3 (D483N), E1 (P397L) and E3 (Q19R) were identified among CACV. Acquisition of the nsP3-483N mutation in Compostela Valley followed by E1-397L/E3-19R in Laguna preceded the nationwide spread in the Philippines. The China isolates possessed two of the amino acid substitutions, nsP3-D483N and E1-P397L whereas the Micronesian and Caribbean CHIKV inherited all the three amino acid substitutions. The unique amino acid substitutions observed among the isolates suggest multiple independent virus dissemination events. The possible biological importance of the specific genetic signatures associated with the rapid global of the virus is not known and warrant future in-depth study and epidemiological follow-up. Molecular evidence, however, supports the Philippines outbreak as the possible origin of the CACV.
Bacterial communities in Haemaphysalis, Dermacentor and Amblyomma ticks collected from wild boar of an Orang Asli Community in Malaysia

Lim FS, Khoo JJ, Tan KK, Zainal N, Loong SK, Khor CS, et al.

Ticks Tick Borne Dis, 2020 03;11(2):101352.
PMID: 31866439 DOI: 10.1016/j.ttbdis.2019.101352

Ticks are hematophagous vectors of arthropod-borne disease agents globally. In Malaysia, despite seroprevalence studies indicating the presence of tick-borne diseases among the indigenous people, the etiological agents of these diseases are still unclear. These indigenous people, also known as the Orang Asli, still live in forested areas with frequent contact with wildlife. Wild boar are ubiquitously found in the forested areas where the Orang Asli communities are located and are commonly hunted as a food supplement. In this study, we aim to determine the tick species parasitizing wild boar from an Orang Asli community, and explore the tick-associated bacterial communities using 16 s rRNA amplicon sequencing on the Ion Torrent PGM™ platform. A total of 72 ticks were collected from three wild boar and were morphologically identified as Haemaphysalis hystricis (n = 32), Dermacentor compactus (n = 15), Amblyomma testudinarium (n = 13), Dermacentor steini (n = 10) and Dermacentor atrosignatus (n = 2). Across all tick samples, 910 bacterial taxa were identified. Although the bacterial communities were not significantly distinct between tick species in beta-diversity analyses, Coxiella, Rickettsia and Francisella were detected at high relative abundance in H. hystricis, D. compactus and D. steini respectively. Many other bacterial genera, including those that have been described in many different tick species, were also identified, including Pseudomonas, Acinetobacter, Staphylococcus and Corynebacterium. Beta-diversity analyses also showed that the bacterial communities were separated based on the animal host from which the ticks were collected from, suggesting that the bacterial communities here may be influenced by the animal skin microflora, host blood or the environment. PCR screening confirmed the presence of Rickettsia sp. related to spotted fever group Rickettsia in some of the ticks. This study provides baseline knowledge of the microbiome of H. hystricis, D. atrosignatus, D. compactus, D. steini and A. testudinarium parasitizing wild boar in this region. The information gained in this study provides the basis to target our efforts in H. hystricis, D. compactus and D. steini for the future investigation of vector competence and the zoonotic potential for the Coxiella, Rickettsia and Francisella detected here, as well as their implications for the risks of tick-borne diseases among the Orang Asli communities.
Fulltext Syringocystadenoma papilliferum arising in a naevus sebaceous

Faheem NAA, Kwan Z, Yong ASW, Ch'ng CC, Tan KK, Naicker M, et al.

Malays J Pathol, 2019 Apr;41(1):47-49.
PMID: 31025637

Naevus sebaceus is a cutaneous hamartoma with the potential of developing into benign or malignant neoplasms. Syringocystadenoma papilliferum (SCAP) have been reported to originate from naevus sebaceus. SCAP is a rare, benign adnexal skin tumour of apocrine or eccrine type of differentiation which typically presents as a nodule or a plaque on the scalp or face. We report a case of syringocystadenoma papilliferum arising in an undiagnosed pre-existing naevus sebaceus in a 56-year-old female.
Vector competence of Malaysian Aedes albopictus with and without Wolbachia to four dengue virus serotypes

Joanne S, Vythilingam I, Teoh BT, Leong CS, Tan KK, Wong ML, et al.

Trop Med Int Health, 2017 09;22(9):1154-1165.
PMID: 28653334 DOI: 10.1111/tmi.12918

OBJECTIVE: To determine the susceptibility status of Aedes albopictus with and without Wolbachia to the four dengue virus serotypes.
METHODS: Two newly colonised colonies of Ae. albopictus from the wild were used for the study. One colony was naturally infected with Wolbachia while in the other Wolbachia was removed by tetracycline treatment. Both colonies were orally infected with dengue virus-infected fresh blood meal. Dengue virus load was measured using quantitative RT-PCR at four-time intervals in the salivary glands, midguts and ovaries.
RESULTS: Wolbachia did not significantly affect Malaysian Ae. albopictus dengue infection or the dissemination rate for all four dengue virus serotypes. Malaysian Ae. albopictus had the highest replication kinetics for DENV-1 and the highest salivary gland and midgut infection rate for DENV-4.
CONCLUSION: Wolbachia, which naturally exists in Malaysian Ae. albopictus, does not significantly affect dengue virus replication. Malaysian Ae. albopictus is susceptible to dengue virus infections and capable of transmitting dengue virus, especially DENV-1 and DENV-4. Removal of Wolbachia from Malaysian Ae. albopictus would not reduce their susceptibility status.
Fulltext Estimating the population health and economic impacts of introducing a pneumococcal conjugate vaccine in Malaysia- an economic evaluation

Shafie AA, Ahmad N, Naidoo J, Foo CY, Wong C, Pugh S, et al.

Hum Vaccin Immunother, 2020 07 02;16(7):1719-1727.
PMID: 31951782 DOI: 10.1080/21645515.2019.1701911

Pneumococcal disease is a potentially fatal bacterial infection that is vaccine-preventable. Malaysia has yet to adopt a pneumococcal conjugate vaccine (PCV) into its national immunization program (NIP). In 2016, pneumonia was the 3rd leading cause of death in children under five in Malaysia, accounting for 3.8% of under-five deaths. Introducing a pneumococcal conjugate vaccine (PCV) is an effective strategy to reduce the disease burden. This study used a decision-analytic model to assess the potential impacts of introducing the available PCVs (13-valent and 10-valent) in Malaysia. Epidemiological and costs inputs were sourced from published literature. For each vaccination program, health outcomes and associated healthcare costs were estimated. The scenarios of initiating PCV13 vs. PCV10 and the status quo (no pneumococcal vaccine) were compared. Serotype trends of Finland and the U.K. were used to model the clinical impacts of PCV10 and PCV13 respectively. The base-case analysis used a societal perspective over a 5-year time horizon. Compared with PCV10, PCV13 was projected to avert an additional 190,628 cases of pneumococcal disease and 1126 cases of death. The acquisition of PCV13 was estimated to cost an incremental US$89,904,777, offset by a cost reduction of -US$250,219,914 on pneumococcal disease-related medical care and lost productivity. PCV13 demonstrated a higher cost-saving potential over PCV10. Compared with no vaccination, PCV13 was estimated as cost-saving. Results were robust across a series of sensitivity analyses. The introduction of PCV13 in a NIP was estimated to reduce a significant burden of disease and to be a cost-saving for the Malaysian health system.
Epidemiology, clinical characteristics, laboratory findings and severity of respiratory syncytial virus acute lower respiratory infection in Malaysian children, 2008-2013

Ng KF, Tan KK, Sam ZH, Ting GS, Gan WY

J Paediatr Child Health, 2017 Apr;53(4):399-407.
PMID: 27704652 DOI: 10.1111/jpc.13375

AIM: The aim of this study is to describe epidemiology, clinical features, laboratory data and severity of respiratory syncytial virus (RSV) acute lower respiratory infection (ALRI) in Malaysian children and to determine risk factors associated with prolonged hospital stay, paediatric intensive care unit (PICU) admission and mortality.
METHODS: Retrospective data on demographics, clinical presentation, outcomes and laboratory findings of 450 children admitted into Tuanku Jaafar Hospital in Seremban, Malaysia from 2008 to 2013 with documented diagnosis of RSV ALRI were collected and analysed.
RESULTS: Most admissions were children below 2 years old (85.8%; 386/450). Commonest symptoms were fever (84.2%; 379/450), cough (97.8%; 440/450) and rhinorrhea (83.6%; 376/450). The median age among febrile patients (n = 379) was 9.0 months with interquartile range (IQR) of 4.0-19.0 months whereas the median age among those who were apyrexial (n = 71) was 2 months with IQR of 1-6 months (P-value <0.001). 15.3% (69/450) needed intensive care and 1.6% (7/450) died. Young age, history of prematurity, chronic comorbidity and thrombocytosis were significantly associated with prolonged hospital stay, PICU admission and mortality.
CONCLUSIONS: Infants less than 6 months old with RSV ALRI tend to be afebrile at presentation. Younger age, history of prematurity, chronic comorbidity and thrombocytosis are predictors of severe RSV ALRI among Malaysian children. Case fatality rate for Malaysian children below 5 years of age with RSV ALRI in our centre is higher than what is seen in developed countries, suggesting that there is room for improvement.
Fulltext Emotional exhaustion is associated with work related stressors: a cross-sectional multicenter study in Malaysian public hospitals

Khoo EJ, Aldubai S, Ganasegeran K, Lee BX, Zakaria NA, Tan KK

Arch Argent Pediatr, 2017 06 01;115(3):212-219.
PMID: 28504485 DOI: 10.5546/aap.2017.eng.212

INTRODUCTION: Emotional exhaustion is an important component of burnout. Burnout is common among doctors. It affects the physical and mental health of doctors, their performance and the quality of care they provide. This study aimed to investigate the level and factors associated with emotional exhaustion among doctors in pediatric practice in Malaysia.
POPULATION AND METHODS: A self-administered questionnaire was used in this multicenter cross-sectional study. It included questions on the socio-demographics, work characteristics, Emotional Exhaustion, Perceived Stress Scale and sources of job stress. Descriptive, univariate and multivariate analysis were conducted using the SPSS software.
RESULTS: A total of 197 doctors working in the Pediatric department in eight hospitals returned complete questionnaire. High and moderate emotional exhaustion was reported by 25.4% and 24.4% doctors, respectively. In bivariate analysis, 29 out of the 38 items of sources of stress showed significant association with emotional exhaustion (p <0.05).The significant predictors of emotional exhaustion in the multivariate analysis were: scoring higher on the Perceived Stress Score, dealing with patient's psychosocial problems, disrespectful interactions with colleagues/ subordinates, lack of appreciation from supervisors, lack of incentives and promotions, time pressures and deadlines to meet, and setting unrealistic goals of having them imposed on oneself (p <0.05). The most common source of stress was dealing with difficult parents (80.2%).
CONCLUSIONS: Emotional exhaustion is associated with sources of stress in the workplace but not with socio-demographic factors.
Sera of patients with systemic lupus erythematosus cross-neutralizes dengue viruses

Zainal N, Tan KK, Johari J, Hussein H, Wan Musa WR, Hassan J, et al.

Microbiol. Immunol., 2018 Oct;62(10):659-672.
PMID: 30259549 DOI: 10.1111/1348-0421.12652

Dengue is the most prevalent mosquito-borne disease in Southeast Asia, where the incidence of systemic lupus erythematosus (SLE) is approximately 30 to 53 per 100,000. Severe dengue, however, is rarely reported among individuals with SLE. Here, whether sera of patients with SLE cross-neutralize dengue virus (DENV) was investigated. Serum samples were obtained from individuals with SLE who were dengue IgG and IgM serology negative. Neutralization assays were performed against the three major DENV serotypes. Of the dengue serology negative sera of individuals with SLE, 60%, 61% and 52% of the sera at 1/320 dilution showed more than 50% inhibition against dengue type-1 virus (DENV-1), DENV-2 and DENV-3, respectively. The neutralizing capacity of the sera was significantly greater against DENV-1 (P
Omeprazole 40 mg o.m. combined with amoxycillin alone or with amoxycillin and metronidazole in the eradication of Helicobacter pylori

Goh KL, Peh SC, Parasakthi N, Wong NW, Tan KK, Lo YL

Am J Gastroenterol, 1994 Oct;89(10):1789-92.
PMID: 7942668

OBJECTIVES: Our objectives were to determine the effect of dual therapy with omeprazole and amoxicillin and of triple therapy with omeprazole, amoxicillin, and metronidazole in the eradication of Helicobacter pylori (HP) and to study the long-term results of eradication in these patients.
METHODS: A prospective, randomized, controlled trial was performed. Patients who were recruited had unequivocal evidence of HP infection based on culture, histology, rapid urease test, and Gram's stain of a tissue smear. Eradication was defined as the absence of bacteria in all tests performed on both corpus and antral biopsies.
RESULTS: The infection was eradicated in 15 of 19 (78.9%) patients randomized to receive dual therapy and in 19 of 22 (86.4%) patients who received triple therapy. We followed the course of 30 patients in whom HP had been eradicated for a prolonged term (up to 12 months). All remained clear of HP. Twenty-five of 28 patients (89.3%) with duodenal ulcers in whom HP was successfully eradicated remained healed at 12 months. Fewer side effects were reported among patients who received the dual therapy.
CONCLUSIONS: Combination therapy with omeprazole and amoxicillin with or without metronidazole is effective in the eradication of HP. In particular, the dual therapy regimen with amoxicillin is not only effective but is also well tolerated by patients.
Hepatic Sarcoidosis: Diagnostic approach and management

Hon SA, Lee LT, Tan KK, Lee RA, Low QJ

Med J Malaysia, 2021 Nov;76(6):914-917.
PMID: 34806684

Sarcoidosis is a multi-systemic, non-caseating granulomatous disorder with an idiopathic aetiology. We report a 58-year-old Malay woman, with underlying type II diabetes mellitus, hypertension and history of stage II pulmonary sarcoidosis presenting with incidental finding of multiple hypodense liver lesions. Her recent contrasted enhanced computed tomography of the abdomen and pelvis demonstrated multiple intra-abdominal lymphadenopathies with evidence of liver and splenic infiltrations. Her ageappropriate malignancy screening was negative while liver biopsy showed non-caseating granulomatous hepatitis consistent with hepatic sarcoidosis. In view of her normal liver enzymes and normalised serum calcium levels, no immunosuppressive therapy was commenced. She remains asymptomatic and is currently under our close monitoring.
Fulltext Small intestinal absorption studies and biopsy findings in Asians, in Singapore

Jayaratnam FJ, Tan KK, Jacob E, Seah CS, Kho KM

Singapore Med J, 1970 Dec;11(4):275-82.
PMID: 5511860

Investigations in 30 healthy control subjects of Indian, Chinese and Malay ethnic origin, revealed that all the Chinese and about two thirds of the Indians and Malays had a normal capacity to absorb D—xylose, Vitamin A, Co58 labelled Vitamin B12 and dietary fat. About a third of the Indians and Malays were unable to absorb one or two of the four test substances used in the absorption studies. Jejunal biopsies did not differ in the 3 ethnic groups and were normal or mildly abnormal. 27 patients presenting with anorexia, a sore tongue and pallor were also investigated. Indians formed the majority of the patients. Diarrhoea occurred in only 51 % of the patients. All had a megaloblastic anaemia. Absorption studies revealed malabsorption of xylose, Vitamin A and Vitamin B12 in the majority but steatorrhoea occurred in only 26% of the patients. Jejunal biopsies were mildly abnormal in 8% and moderately or severely abnormal in 92 % of the patients. All responded to folic acid or Vitamin B12 therapy. 16 patients were restudied after 5 to 24 months therapy and the majority were found to have improved. Results of investigations and response to therapy indicate that these patients were suffering from tropical sprue. These studies indicate that tropical sprue in Singapore affects Indians mainly and can often present without diarrhoea and steatorrhoea.
Fulltext Draft genome of the emerging pathogen, Kocuria marina, isolated from a wild urban rat

Loong SK, Tan KK, Zainal N, Phoon WH, Zain SNM, AbuBakar S

Mem Inst Oswaldo Cruz, 2017 Dec;112(12):857-859.
PMID: 29211248 DOI: 10.1590/0074-02760170132

Kocuria marina has recently emerged as a cause for catheter-related bloodstream infections in patients with underlying health complications. One K. marina strain was recently isolated from the lung tissues of a wild urban rat (Rattus rattus diardii) caught during rodent surveillance. Here, we present the draft genome of the first K. marina animal isolate, K. marina TRE150902.
Fulltext Pernicious anaemia among Asians in Singapore

Jayaratnam FJ, Seah CS, Da Costa JL, Tan KK, O'Brien W

Br Med J, 1967 Jul 01;3(5556):18-20.
PMID: 6027375
Fulltext Non-classical Bordetella sp. (closely related to Bordetella hinzii and Bordetella pseudohinzii) lower respiratory tract infection in a patient with extensive bronchiectasis: a case report

Loong SK, Liam CK, Karunakaran R, Tan KK, Mahfodz NH, AbuBakar S

J Int Med Res, 2024 Jan;52(1):3000605231214464.
PMID: 38216150 DOI: 10.1177/03000605231214464

An increasing number of reports have described the pathogenic nature of several non-classical Bordetella spp. Among them, Bordetella hinzii and Bordetella pseudohinzii have been implicated in a myriad of respiratory-associated infections in humans and animals. We report the isolation of a genetically close relative of B. hinzii and B. pseudohinzii from the sputum of a woman in her early 60s with extensive bronchiectasis who presented with fever and brown colored sputum. The isolate had initially been identified as Bordetella avium by API 20NE, the identification system for non-enteric Gram-negative rod bacteria. Sequencing of the 16S rDNA, ompA, nrdA, and genes used in the Bordetella multilocus sequence typing scheme could not resolve the identity of this Bordetella isolate. Whole-genome single nucleotide polymorphism analysis positioned the isolate between B. hinzii and B. pseudohinzii in the phylogenetic tree, forming a distinct cluster. Whole-genome sequencing enabled the further identification of this rare organism, and should be considered for wider applications, especially the confirmation of organism identity in the clinical diagnostic microbiology laboratory.
Fulltext Repair of segmental load-bearing bone defect by autologous mesenchymal stem cells and plasma-derived fibrin impregnated ceramic block results in early recovery of limb function

Ng MH, Duski S, Tan KK, Yusof MR, Low KC, Rose IM, et al.

Biomed Res Int, 2014;2014:345910.
PMID: 25165699 DOI: 10.1155/2014/345910

Calcium phosphate-based bone substitutes have not been used to repair load-bearing bone defects due to their weak mechanical property. In this study, we reevaluated the functional outcomes of combining ceramic block with osteogenic-induced mesenchymal stem cells and platelet-rich plasma (TEB) to repair critical-sized segmental tibial defect. Comparisons were made with fresh marrow-impregnated ceramic block (MIC) and partially demineralized allogeneic bone block (ALLO). Six New Zealand White female rabbits were used in each study group and three rabbits with no implants were used as negative controls. By Day 90, 4/6 rabbits in TEB group and 2/6 in ALLO and MIC groups resumed normal gait pattern. Union was achieved significantly faster in TEB group with a radiological score of 4.50 ± 0.78 versus ALLO (1.06 ± 0.32), MIC (1.28 ± 0.24), and negative controls (0). Histologically, TEB group scored the highest percentage of new bone (82% ± 5.1%) compared to ALLO (5% ± 2.5%) and MIC (26% ± 5.2%). Biomechanically, TEB-treated tibiae achieved the highest compressive strength (43.50 ± 12.72 MPa) compared to those treated with ALLO (15.15 ± 3.57 MPa) and MIC (23.28 ± 6.14 MPa). In conclusion, TEB can repair critical-sized segmental load-bearing bone defects and restore limb function.
Fulltext Detection of dengue viruses using reverse transcription-loop-mediated isothermal amplification

Teoh BT, Sam SS, Tan KK, Johari J, Danlami MB, Hooi PS, et al.

BMC Infect Dis, 2013;13:387.
PMID: 23964963 DOI: 10.1186/1471-2334-13-387

BACKGROUND: Early and rapid detection of dengue virus (DENV) infection during the febrile period is crucial for proper patient management and prevention of disease spread. An easy to perform and highly sensitive method is needed for routine implementation especially in the resource-limited rural healthcare settings where dengue is endemic.
METHODS: A single-tube reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay with a set of nine primers was developed for the detection of all four DENV serotypes and their different genotypes. The sensitivity and specificity of the RT-LAMP were evaluated. The clinical applicability of RT-LAMP assay for detection of DENV RNA was assessed in a total of 305 sera of clinically-suspected dengue patients. The test results of RT-LAMP were statistically compared to those of quantitative reverse transcription-polymerase chain reaction (qRT-PCR), IgM- and IgG-capture enzyme-linked immunosorbent assays (ELISA).
RESULTS: Acute DENV infection was confirmed in 171 samples (n = 305); 43.3% (74/171) and 46.8% (80/171) of the samples were positive for DENV using RT-LAMP and qRT-PCR, respectively. The combination of RT-LAMP with the dengue IgM and IgG ELISA increased detection of acute DENV infection to 97.7% (167/171), in comparison to only 70.8% (121/171) when dengue IgM and IgG ELISA alone were used. The RT-LAMP assays showed high concordance (κ = 0.939) with the qRT-PCR. The RT-LAMP assay detected up to 10 copies of virus RNA within an hour but 100% reproducibility (12/12) was achieved with 100 copies. There was no cross reactivity of RT-LAMP with other closely related arboviruses.
CONCLUSION: The RT-LAMP assay developed in this study is sensitive, specific and simple to perform. The assay improved the detection of dengue when used in combination with serological methods.
Differential osteogenic activity of osteoprogenitor cells on HA and TCP/HA scaffold of tissue engineered bone

Ng AM, Tan KK, Phang MY, Aziyati O, Tan GH, Isa MR, et al.

J Biomed Mater Res A, 2008 May;85(2):301-12.
PMID: 17688285

Biomaterial, an essential component of tissue engineering, serves as a scaffold for cell attachment, proliferation, and differentiation; provides the three dimensional (3D) structure and, in some applications, the mechanical strength required for the engineered tissue. Both synthetic and naturally occurring calcium phosphate based biomaterial have been used as bone fillers or bone extenders in orthopedic and reconstructive surgeries. This study aims to evaluate two popular calcium phosphate based biomaterial i.e., hydroxyapatite (HA) and tricalcium phosphate/hydroxyapatite (TCP/HA) granules as scaffold materials in bone tissue engineering. In our strategy for constructing tissue engineered bone, human osteoprogenitor cells derived from periosteum were incorporated with human plasma-derived fibrin and seeded onto HA or TCP/HA forming 3D tissue constructs and further maintained in osteogenic medium for 4 weeks to induce osteogenic differentiation. Constructs were subsequently implanted intramuscularly in nude mice for 8 weeks after which mice were euthanized and constructs harvested for evaluation. The differential cell response to the biomaterial (HA or TCP/HA) adopted as scaffold was illustrated by the histology of undecalcified constructs and evaluation using SEM and TEM. Both HA and TCP/HA constructs showed evidence of cell proliferation, calcium deposition, and collagen bundle formation albeit lesser in the former. Our findings demonstrated that TCP/HA is superior between the two in early bone formation and hence is the scaffold material of choice in bone tissue engineering.
Fulltext Bone graft substitute using hydroxyapatite scaffold seeded with tissue engineered autologous osteoprogenitor cells in spinal fusion: early result in a sheep model

Tan KK, Tan GH, Shamsul BS, Chua KH, Ng MHA, Ruszymah BHI, et al.

Med J Malaysia, 2005 Jul;60 Suppl C:53-8.
PMID: 16381285

Spinal fusion using autologous bone graft is performed in an increasing rate for many spinal disorders. However, graft harvesting procedure is associated with prolonged operation time and potential donor site morbidity. We produced an engineered 'bone graft' substitute by using porous hydroxyapatite (HA) scaffold seeded with autologous bone marrow osteoprogenitor cells (OPCs) and fibrin. This obviates bone graft harvesting, thus eliminates donor site morbidity and shortens the operation time. The aim of this study is to evaluate Hydroxyapatite (HA) ceramics as scaffold for autologous tissue engineered bone construct for spinal fusion in a sheep model. The sheep's marrow was aspirated from iliac crest. The bone marrow mesenchymal stem cells (BMMSCs) were cultured for several passages in the presence of growth and differentiation factors to increase the number of OPCs. After the cultures reached confluence, they were trypsinized and seeded on Hydroxyapatite scaffold (HA). Approximately 5 million cells were generated after 3 weeks of culture. Microscopically, very tight Colony Forming Units (CFU-Fs) were seen on monolayer culture. The Von Kossa and Alizarin Red staining of monolayer culture showed positive mineralization areas; indicating the presence of OPCs. Sheep underwent a posterolateral spinal fusion in which scaffolds with or without OPCs seeded were implanted on both sides of the lumbar spine (L1-L2). Intended fusion segments were immobilized using wires. At the end of third month, the fusion constructs were harvested for histological examination. Fibrous tissue infiltration found in the inter-connecting pores of plain HA ceramics indicates inefficient new bone regeneration. New bone was found surrounding the HA ceramics seeded with autologous cells. The new bone is probably formed by the sheep BMMSCs that were initially encapsulating HA while it remained intact. The new bone is naturally fused with the vertebrae. In conclusion, the incorporation of autologous bone marrow cells improved the effectiveness of HA ceramics as 'bone graft' substitute for spinal fusion.
Comparison of bioengineered human bone construct from four sources of osteogenic cells

Ng AM, Saim AB, Tan KK, Tan GH, Mokhtar SA, Rose IM, et al.

J Orthop Sci, 2005;10(2):192-9.
PMID: 15815868

Osteoprogenitor cells have been reported to be present in periosteum, cancellous and cortical bone, and bone marrow; but no attempt to identify the best cell source for bone tissue engineering has yet been reported. In this study, we aimed to investigate the growth and differentiation pattern of cells derived from these four sources in terms of cell doubling time and expression of osteoblast-specific markers in both monolayer cells and three-dimensional cell constructs in vitro. In parallel, human plasma derived-fibrin was evaluated for use as biomaterial when forming three-dimensional bone constructs. Our findings showed osteoprogenitor cells derived from periosteum to be most proliferative followed by cortical bone, cancellous bone, and then bone marrow aspirate. Bone-forming activity was observed in constructs formed with cells derived from periosteum, whereas calcium deposition was seen throughout the constructs formed with cells derived from cancellous and cortical bones. Although no mineralization activity was seen in constructs formed with osteoprogenitor cells derived from bone marrow, well-organized lacunae as would appear in the early phase of bone reconstruction were noted. Scanning electron microscopy evaluation showed cell proliferation throughout the fibrin matrix, suggesting the possible application of human fibrin as the bioengineered tissue scaffold at non-load-bearing sites.

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