Sterilization of HIV with irradiation: relevance to infected bone allografts.

BACKGROUND: Bone allograft banks commonly sterilize frozen bone by irradiation. The dose-response relationship for HIV is calculated and the dose required to inactivate the bioburden of virus that may be present in allograft bone is determined. METHODS: A virus titre experiment is performed using irradiated frozen HIV. The virus is maintained on dry ice (approximately -70 degrees C) and is exposed to a cobalt 60 source with 0-40 kGy irradiation at 5 kGy intervals. Lymphocyte cell cultures are exposed to serial dilutions of the irradiated virus. The virus titre is quantified by cytological changes of HIV infection and p24 immunofluorescence. RESULTS: There is a linear relationship between the virus titre and the radiation dose delivered. The inactivation rate of irradiated virus was 0.1134 log10 tissue culture infective doses 50/mL per kGy (95% confidence intervals, 0.1248-0.1020). The irradiation dose required to inactivate the HIV bioburden in allograft bone is 35 kGy. The irradiation dose required to achieve a sterility assurance level of 10(-6) is 89 kGy. This dose exceeds current recommendations for sterilizing medical products and the current practice of many bone banks. CONCLUSIONS: It is concluded that gamma irradiation should be disregarded as a significant virus inactivation method for bone allografts.     

STERILIZATION OF HIV WITH IRRADIATION: RELEVANCE TO INFECTED BONE ALLOGRAFTS

Background : Bone allograft banks commonly sterilize frozen bone by irradiation. The dose–response relationship for HIV is calculated and the dose required to inactivate the bioburden of virus that may be present in allograft bone is determined. Methods : A virus titre experiment is performed using irradiated frozen HIV. The virus is maintained on dry ice (approximately –70°C) and is exposed to a cobalt 60 source with 0–40 kGy irradiation at 5 kGy intervals. Lymphocyte cell cultures are exposed to serial dilutions of the irradiated virus. The virus titre is quantified by cytological changes of HIV infection and p24 immunofluorescence. Results : There is a linear relationship between the virus titre and the radiation dose delivered. The inactivation rate of irradiated virus was 0.1134 log₁₀ tissue culture infective doses ₅₀/mL per kGy (95% confidence intervals, 0.1248–0.1020). The irradiation dose required to inactivate the HIV bioburden in allograft bone is 35 kGy. The irradiation dose required to achieve a sterility assurance level of 10^–6 is 89 kGy. This dose exceeds current recommendations for sterilizing medical products and the current practice of many bone banks. Conclusions : It is concluded that gamma irradiation should be disregarded as a significant virus inactivation method for bone allografts.     

Demineralization for inactivation of infectious retrovirus in systemically infected cortical bone: in vitro and in vivo experimental studies

BACKGROUND: Clinical and experimental studies have demonstrated viral transmission through the transplantation of fresh-frozen infected bone. While sterilization methods sufficient to inactivate the human immunodeficiency virus (HIV) have been shown to markedly alter osteoconductive and osteoinductive properties of bone allografts, the ability of a process for creating demineralized bone matrix to abrogate transmission of a retrovirus has not been investigated, to our knowledge. We hypothesized that a clinically accepted demineralization procedure would alter the nucleic acids of the feline leukemia virus (FeLV, a retrovirus with a structure and replication cycle similar to those of HIV), inactivating the virus in infected bone and rendering it noninfectious. METHODS: Bone infected with FeLV was demineralized with a method employed for creating demineralized bone matrix powder. The effects of demineralization on cellular and (pro)viral nucleic acids were examined with use of gel electrophoresis and quantitative polymerase chain reaction, respectively. To compare the infectivity of the demineralized bone matrix with that of mineralized bone particles in cell cultures and in animals in which they had been implanted, we measured FeLV p27 antigen and (pro)viral nucleic acids as well as antiviral antibodies. RESULTS: Demineralization of FeLV-infected bone appeared to inactivate the virus by degradation and fragmentation of the DNA, rendering it noninfectious in both in vitro and in vivo test systems. In contrast, untreated mineralized FeLV-infected bone contained intact nucleic acids and readily transmitted the virus in both test systems. CONCLUSIONS: The demineralization process inactivated infectious retrovirus in infected cortical bone, thereby preventing disease transmission.     

Modulation of cell phenotype in human osteoblast-like cells by the simian virus 40

At present, the majority of in vitro research into bone metabolism is performed on either primary cultures of bone or osteosarcoma lines. A better model of the behaviour of normal bone cells would be a cell line derived from normal, adult bone that retained osteoblast-like characteristics. We infected a culture of bone cells from adult humans with simian virus 1613, a variant of the simian virus 40, and obtained 12 clones of variable morphology. The clones were maintained in culture for as long as 6 months. Population doubling times, synthesis of alkaline phosphatase and osteocalcin, secretion of mineral, morphology, and ability to withstand freezing were examined. SV/EC cell morphology varied from the polygonal, osteoblast-like to the bipolar, fibroblast-like. Population doubling times ranged from 0.55 to 2.8 days (compared with 3.9 days for the nontransformed human osteoblast-like cells). Synthesis of alkaline phosphatase varied but was less than that by the human osteoblast-like cells. With the exception of clone 11, all of the transformed clones synthesised mineral in vitro under mineralising conditions. Four clones showed increased synthesis of alkaline phosphatase and increased population doubling times after passaging. All of the clones were successfully frozen and thawed, but, unlike normal human osteoblast-like cells, none responded to stimulation with parathyroid hormone. These results demonstrate that normal human bone cells can incorporate the simian virus 1613 while retaining some of their osteoblast-like characteristics. Such manipulation affords these cells an extended lifespan, and this, together with their ability to withstand freezing, makes them a potentially useful tool in bone research.     

Fracture healing in HIV-positive populations

Highly active anti-retroviral therapy has transformed HIV into a chronic disease with a long-term asymptomatic phase. As a result, emphasis is shifting to other effects of the virus, aside from immunosuppression and mortality. We have reviewed the current evidence for an association between HIV infection and poor fracture healing. The increased prevalence of osteoporosis and fragility fractures in HIV patients is well recognised. The suggestion that this may be purely as a result of highly active anti-retroviral therapy has been largely rejected. Apart from directly impeding cellular function in bone remodelling, HIV infection is known to cause derangement in the levels of those cytokines involved in fracture healing (particularly tumour necrosis factor-alpha) and appears to impair the blood supply of bone. Many other factors complicate this issue, including a reduced body mass index, suboptimal nutrition, the effects of anti-retroviral drugs and the avoidance of operative intervention because of high rates of wound infection. However, there are sound molecular and biochemical hypotheses for a direct relationship between HIV infection and impaired fracture healing, and the rewards for further knowledge in this area are extensive in terms of optimised fracture management, reduced patient morbidity and educated resource allocation. Further investigation in this area is overdue.     

The Risk of HIV, HBV, HCV and HTLV Infection Among Musculoskeletal Tissue Donors in Australia

In Australia, there are no current national estimates of the risks of transmission of human immunodeficiency virus (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV) or human T-lymphotrophic virus (HTLV) by musculoskeletal tissue transplantation. We determined the prevalence rates of antibodies against HIV (anti-HIV), HCV (anti-HCV) and HTLV (anti-HTLV) and Hepatitis B surface antigen (HBsAg) for 12,415 musculoskeletal tissue donors from three major bone tissue banks across Australia for the period 1993-2004. The prevalence (per 100,000 persons) was 64.44 for anti-HIV, 407.13 for HBsAg, 534.63 for anti-HCV and 121.88 for anti-HTLV. The estimated probability of viremia at the time of donation was 1 in 128,000, 1 in 189,000, 1 in 55,000 and 1 in 118,000, respectively. With the addition of nucleic acid amplification testing (NAT), the probability of donor viremia would be reduced to 1 in 315,000 for HIV, 1 in 385,000 for HBV and 1 in 500,000 for HCV. The prevalence of HIV, HBV, HCV and HTLV although low, are higher among musculoskeletal tissue donors than among first-time blood donors. The risks associated with musculoskeletal donation will be reduced with NAT, though further cost analysis is required prior to its implementation.     

Inactivation of HIV by application of heat and radiation: implication in bone banking with irradiated allograft bone

We developed methods for inactivating the human immunodeficiency virus by heat and ionizing radiation and tested the effects of these treatments on the mechanical strength of bone. Simultaneous use of heat and radiation caused a considerably greater inactivation of HIV than the additive effects of the two separate treatments, but also caused a significant reduction in the maximum load sustained by the bone specimens tested with an Instrom machine. Application of the same doses but given in the sequential fashion of radiation followed by heat also caused marked inactivation of HIV and had less effect on the mechanical strength of the bone.     

Bone transplantation and human immunodeficiency virus. An estimate of risk of acquired immunodeficiency syndrome (AIDS)

The possibility of transplanting a bone allograft from a donor infected with human immunodeficiency virus (HIV) is remote, provided there is a combination of rigorous donor selection and exclusion, screening for the HIV antigen and antibody, and histopathologic studies of donor tissues. The chance of obtaining a bone allograft from an HIV-infected donor who failed to be excluded by the above techniques is calculated to be one in well over a million, using average estimates. On the other hand, if adequate precautions are not taken (for example, by testing only for antibodies to HIV), the risk might be as high as one in 161.     

Inactivation of HIV by application of heat and radiation: Implication in bone banking with irradiated allograft bone

We developed methods for inactivating the human immunodeficiency virus by heat and ionizing radiation and tested the effects of these treatments on the mechanical strength of bone. Simultaneous use of heat and radiation caused a considerably greater inactivation of HIV than the additive effects of the two separate treatments, but also caused a significant reduction in the maximum load sustained by the bone specimens tested with an Instrom machine. Application of the same doses but given in the sequential fashion of radiation followed by heat also caused marked inactivation of HIV and had less effect on the mechanical strength of the bone.     

HIV Infection and Osteoporosis: Pathophysiology, Diagnosis, and Treatment Options

As the population with HIV continues to age, specialists in HIV care are increasingly encountering chronic health conditions, which now include osteoporosis, osteopenia, and fragility fractures. The pathophysiology of the bone effects of HIV infection is complex and includes traditional risk factors for bone loss as well as specific effects due to the virus itself, chronic inflammation, and HAART. Examining risk factors for low bone density and screening of certain patients is suggested, and consideration should be given to treatment for those considered high risk for fracture.     

Transfusion transmitted human T-lymphotropic virus infections

Human immunodeficiency virus (HIV) type 1 has emerged during the 1980s as an important transfusion transmitted agent. The impact of HIV depends on the epidemiology and characteristics of the virus, serological response to infection and efficacy of serological tests to identify infected blood units. These factors will be described in the present article. The risk to acquire HIV today, by transfusion of anti- HIV screened blood, is extremely small in Scandinavia and the Western world. The risk can be further minimized by more sensitive screening tests and an improved donor information. The epidemiology in Scandinavia of other members of the human T-lymphotrophic viruses, HTLV-1 and HIV-2, has to be further investigated.     

Sterilization of HIV by gamma irradiation

Summary A human immuno-deficiency virus (HIV) infected bone allograft model has been created using HTLV-IIIB virus in a concentration simulating a massively HIV infected bone allograft donor [HTLV-III is the denomination initially given to the human immuno-deficiency by the american team of Prof. Gallo. It represent the virus HIV 1 of the present international nomenclature]. 5×10⁴ tissue culture infective doses per ml. of virus were placed within the medullary cavity of bovine femora and tibiae with a radiation dosimeter, and the ends sealed with lead. The bone/virus model was maintained at –70° C while being irradiated with 1 to 4 megarads of gamma irradiation in increments of 0.5 megarads. The study showed that the HTLV-IIIB virus is a relatively radio-resistant organism, a property common to most viruses. The results suggest that HTLV-IIIB can be inactivated in bone infected with a clinically significant viral load, as may be found in donors who are initially negative when screened for HIV. It is recommended that bone allografts which are secondarily sterilized by gamma irradiation receive at least 2.5 megarads. The amount of radiation absorbed by the bone cortex was minimal.

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Résumé Un modèle d'allogreffe osseuse infectée par le virus de l'immuno-déficience humaine (VIH) a été créé en utilisant le virus HTLV-III B [HTLV-III est l'appellation donnée initialement au virus de l'immuno-déficience humaine par l'équipe américaine du Pr Gallo. Il équivaut donc au virus HIV 1 de l'actuelle nomenclature internationale.] à une concentration simulant le cas le plus grave: correspondant à un donneur d'allogreffe osseuse infecté de façon massive par le VIH, 5×10⁴ doses de culture de tissus infectés par ml de virus ont été placées dans le canal médullaire d'un fémur et d'un tibia bovins avec un dosimètre à radiations, les extrémités du modèle étant scellées par du plomb. Le modèle os/virus a été maintenu à une température de –70° Celsius tout en étant soumis à une irradiation gamma de 1 à 4 mégarads par paliers de 0,5 mégarads. Cette étude a montré que le virus HTLV-III B est un organisme relativement résistant aux radiations, propriété commune à la plupart des virus. Les résultats laissent à penser que le virus HTLV-III B pourrait être inactivé dans un os infecté par une charge virale cliniquement significative, comme on peut en trouver chez les donneurs qui réagissent initialement de façon négative aux tests VIH courants. Il est recommandé que les allogreffes osseuses stérilisées par radiations gamma reçoivent au moins 2,5 mégarads. La quantité de radiation absorbée par la corticale osseuse était minimale.

     

Risk factors for decreased bone density and effects of HIV on bone in the elderly

SUMMARY: Most studies of bone density in HIV-infected individuals focus on young men. This study compares differences in bone density in elderly HIV positive men and women to HIV negative controls. Bone density was lower in the lumbar spine and hip in the HIV-infected group. Antiretrovirals may be associated with decreased bone mineralization. INTRODUCTION: Individuals with human immunodeficiency virus (HIV) may be at increased risk for osteoporosis. Prolonged exposures to HIV and/or antiretroviral therapy are possible causes for this association. This study compares differences in bone mineral density (BMD) in elderly HIV positive men and women to HIV negative controls. METHODS: A cross-sectional study was conducted among 57 HIV-infected and 47 HIV negative subjects over age 55. BMD at the lumbar spine and total hip and markers of bone turnover were compared. RESULTS: BMD was borderline lower in the lumbar spine and significantly lower in the hip in the HIV-infected group. Controlling for age, sex, race and body mass index, differences between the groups were significant at both sites. There was no difference in markers of bone turnover between the groups. Tenofovir use was significantly associated with decreased BMD at the spine while protease inhibitor use was significantly associated with decreased BMD at the hip. CONCLUSION: Elderly men and women with HIV have lower bone mass than HIV negative controls. Decreased body mass index was the most important risk factor associated with decreased BMD. Bone demineralization was observed among HIV-infected subjects receiving either tenofovir or a protease inhibitor.     

Role of testosterone in maintaining lean body mass and bone density in HIV-infected patients

Low testosterone levels are common in both men and women with human immunodeficiency virus (HIV) infection and may contribute to loss of lean body mass and AIDS wasting. Causes of low testosterone levels are complex and may include chronic illness, HIV infection and its complications, medications used to treat HIV and opportunistic diseases, and normal aging-related declines. In the majority of studies addressing the use of testosterone treatment in HIV-infected patients, testosterone has been found to help prevent loss of lean body and muscle mass. Whether the combination of exercise and testosterone is more effective in preventing loss of lean body mass than either therapy alone is not yet clear and warrants further study. In addition to its effects on body composition, testosterone treatment results in improved mood and libido in HIV-infected women and increased bone mineral density in HIV-infected men. Testosterone may thus make a valuable contribution to the treatment of HIV-infected individuals. International Journal of Impotence Research (2003) 15, Suppl 4, S21-S25. doi:10.1038/sj.ijir.3901032     

Allograft safety: viral inactivation with bone demineralization

A study was performed to validate the effectiveness of a bone demineralization process with respect to its inactivation of viruses. The viruses selected for study included human immunodeficiency virus (HIV), duck hepatitis B virus (a model for human hepatitis B), bovine viral diarrheal virus (a model for human hepatitis C), human cytomegalovirus, and human poliovirus (a model for small nonenveloped viruses, e.g., hepatitis A). This study was performed in compliance with Good Laboratory Practice regulations using validation methodology similar to that used to ensure the safety of blood derivatives and other products. Use of the bone demineralization process described in this report resulted in a reduction in infectivity of greater than one million (10(6)) for all viruses and as much as one trillion (10(12)) for the poliovirus.     

HIV and Bone Loss

The use of antiretroviral therapy has significantly reduced the number of deaths due to HIV/AIDS. However, no current therapy can suppress the virus completely, and as the HIV-infected population continues to live longer new complications are emerging from the persistence of the virus and use of antiretroviral therapy. This review summarizes the clinical evidence linking HIV-associated osteoporosis to direct infection and antiretroviral therapy (ART) use. The purported molecular mechanisms involved in bone loss are also reviewed. Additionally, recommendations regarding the pharmacologic management of HIV/ART-related osteoporosis are given.     

Effects of HIV infection and antiretroviral therapy with ritonavir on induction of osteoclast-like cells in postmenopausal women

Summary  

Ritonavir (RTV) is a commonly used antiretroviral associated with bone loss. We show that peripheral blood mononuclear cells (PBMCs) from human immunodeficiency virus (HIV)-positive women on RTV are more likely to differentiate into osteoclast-like cells when cultured with their own sera than PBMCs and sera from HIV− women or HIV+ on other antiretrovirals.     

Moderate heat treatment of bone allografts

Summary The use of bone allografts is often essential in orthopedic surgery. Strict donor screening, including HIV testing 3 months postoperatively, is mandatory before a transplant may be used. Yet these measures do not definitely rule out the possibility of HIV transmission, as there is a window period before infection is revealed by blood testing. Accordingly, there is a need for virus inactivation methods that can be used on bone allografts. As radiation treatment and chemical methods have a number of disadvantages, we chose a moderate heat treatment of 65°C for a series of animal experiments. In 12 rabbit femoral condyles, moderate-heattreated bone allografts were implanted into 6-mm drill holes. Twelve normal allografts and 12 empty drill holes served as controls. Radiologic and histological evaluation up to 12 weeks postoperatively revealed slow spontaneous bone remodeling from the rim to the center of the empty cavities. Normal deep frozen allografts were quickly integrated after a short period of osteoclast reaction around the transplant, with occasional bone bridges between host and allgraft. The examination of heattreated allografts showed no differences to the controls, including morphologic aspects and the time course of osteointegration. Five zones of bone repair and osteointegration were distinguished. We conclude that thermal treatment of bone allografts has no adverse effects on osteointegration in the rabbit femoral condyle. Thus, it may contribute to improving safety in human bone transplantation.     

Rapidly progressive periodontal disease associated with human immunodeficiency virus

Severe periodontal inflammation with generalized dental plaque accumulation, spontaneous and severe gingival bleeding, fungal infection, and interdental papillae necrosis are presented in a patient infected with human immunodeficiency virus (HIV). Bite-wing radiographs revealed a generalized horizontal alveolar bone loss of 7-8 millimetres in both arches. Erythematous patches were noted on the gingival mucosa in both jaws. DNA testing was performed to indentify the periodontopathogens. The patient had no signs or symptoms of acquired immunodeficiency syndrome. This case-report presents the massive periodontal destruction that occurred in a patient infected with HIV. Therefore, it is highly recommended that patients infected with HIV should be regularly monitored to aid in early detection and to provide proper management of periodontal inflammatory conditions to minimize its destruction.