A Human Papillomavirus (HPV) In Vitro Neutralization Assay That Recapitulates the In Vitro Process of Infection Provides a Sensitive Measure of HPV L2 Infection-Inhibiting Antibodies

Papillomavirus L2-based vaccines have generally induced low-level or undetectable neutralizing antibodies in standard in vitro assays yet typically protect well against in vivo experimental challenge in animal models. Herein we document that mice vaccinated with an L2 vaccine comprising a fusion protein of the L2 amino acids 11 to 88 of human papillomavirus type 16 (HPV16), HPV18, HPV1, HPV5, and HPV6 were uniformly protected from cervicovaginal challenge with HPV16 pseudovirus, but neutralizing antibodies against HPV16, -31, -33, -45, or -58 were rarely detected in their sera using a standard in vitro neutralization assay. To address this discrepancy, we developed a neutralization assay based on an in vitro infectivity mechanism that more closely mimics the in vivo infectious process, specifically by spaciotemporally separating primary and secondary receptor engagement and correspondingly by altering the timing of exposure of the dominant L2 cross-neutralizing epitopes to the antibodies. With the new assay, titers in the 100 to 10,000 range were measured for most sera, whereas undetectable neutralizing activities were observed with the standard assay. In vitro neutralizing titers measured in the serum of mice after passive transfer of rabbit L2 immune serum correlated with protection from cervicovaginal challenge of the mice. This “L2-based” in vitro neutralization assay should prove useful in critically evaluating the immunogenicity of L2 vaccine candidates in preclinical studies and future clinical trials.     

An animal model of cortical and callosal pathology in multiple sclerosis

The pathological and radiological hallmarks of multiple sclerosis (MS) include multiple demyelinated lesions disseminated throughout the white matter of the central nervous system (CNS). More recently, the cerebral cortex has been shown to be affected in MS, but the elucidation of events causing cortical demyelination has been hampered by the lack of animal models reflecting such human cortical pathology. In this report, we have described the presence of cortical gray matter and callosal white matter demyelinating lesions in the chronic experimental autoimmune encephalomyelitis (EAE) mouse model of MS. Similar to the pathological lesions of MS patients, EAE lesions have been classified as type I-leukocortical, type II-intracortical and type III-subpial. All of these lesions had varying degrees of demyelination, inflammatory cells and reactive astrocytes. Similar to MS, cortical layers during EAE showed demyelination, microglia activation, synaptic protein alterations and apoptotic cells. In addition, the callosal white matter during EAE had many inflammatory demyelinating lesions and axon degeneration. Functional electrophysiological conduction analysis showed deficits in both myelinated and unmyelinated callosal axons during early and late EAE. The chronic EAE mouse model has features that mimic cortical and callosal pathology of MS, and can be potentially used to screen agents to prevent these features of disease.     

Factors predictive of the perceived osteoporosis–fracture link in fragility fracture patients

Objective

Given the asymptomatic nature of osteoporosis, a fragility fracture provides an opportunity to make the issue of osteoporosis relevant to patients. Patients who link their fragility fracture with osteoporosis are more likely to initiate osteoporosis treatment, yet to date, we know little about who is likely to make this link. This study examined whether demographic, health, and osteoporosis belief factors predicted a perceived link between a fragility fracture and osteoporosis.

Study design

This longitudinal cohort study analyzed baseline and follow up data collected as part of a provincial osteoporosis screening initiative targeting fragility fracture patients. Logistic regression analysis was used to examine the relationship between hypothesized predictors and the outcome.

Main outcome measure

Patient perception of the osteoporosis–fracture link at follow up.

Results

At baseline, 93% (1615/1735) of patients did not believe their fracture could have been caused by osteoporosis. Of these, only 8.2% changed this perception at follow up. Adjusted analyses showed that baseline characteristics associated with making the osteoporosis–fracture link at follow up were: a previous fracture (odds ratio (OR) 1.7, confidence interval (CI) 1.2–2.6), perception of osteoporosis pharmacotherapy benefits OR 1.2 (CI 1.0–1.5), diagnosis of rheumatoid arthritis OR 2.6 (CI 1.4–4.9) and the perception of bones as “thin” OR 8.2 (CI 5.1–13.1).

Conclusion

These results shed more light on patient-level barriers to osteoporosis management following an osteoporosis educational programme. They may be used to identify patients less likely to make the link between their fracture and osteoporosis and to inform interventions for this patient group.     

A Novel System to Study the Impact of Epithelial Barriers on Cellular Metabolism

Medications introduced into the systematic circulation must be transported across biological barriers such as skin, gastrointestinal, or bronchial epithelia, which can alter their kinetic and metabolic profiles. It is, therefore, important to understand diffusion kinetics across barrier membranes when choosing a dosing regime that will elicit the greatest cellular response. An in vitro system that combines membrane transport studies with a downstream cell culture chamber has been developed. The system has been tested with skin and a small intestine model (Caco-2 cell monolayers) as barriers, the peroxovanadium compound [VO(O₂)₂ 1, 10 phenanthroline] bpV(phen), as the test chemical, Hep-G2 (liver) as the test cells, and glucose consumption as the test assay. Peroxovanadium has insulin mimetic properties and has been previously demonstrated to effectively lower blood glucose levels in diabetic rats when administered transdermally. A dose of 10 mM bpV(phen) placed on the skin epidermis with a continuous iontophoretic current of 0.5 mA/cm² for 4.5 h led to a net 22% increase in glucose consumption by Hep-G2 cells. The same dose of bpV(phen) passively diffusing across a Caco-2 cell monolayer led to an increase in glucose consumption by Hep-G2 cells of 23%. This system is highly versatile and can be used to study many other processes, involving a variety of biological membranes, cell types, chemicals and assays, making it a valuable research tool. © 2000 Biomedical Engineering Society. PAC00: 8716Uv, 8715Vv     

Stroke,IL-1ra,IL1RN,Infection and Outcome

Background

Infection is a common phenomenon following stroke, and adversely affects outcome. Previous studies suggest that interleukin-1 receptor antagonist (IL-1ra) and single nucleotide polymorphisms (SNPs) in the IL1RN gene might influence the risk of post-stroke infection and outcome. In this study, we addressed the effects of the rs4251961 SNP in IL1RN on infection risk and outcome.

Methods

Subjects with acute ischemic stroke were enrolled within 72 h of symptom onset and followed up to 1 year. Plasma IL-1ra was measured at multiple time points and outcome assessed at 1, 3, 6, and 12 months. Active surveillance for infection occurred while subjects were hospitalized. Subjects were genotyped for the IL1RN rs4251961 polymorphism.

Results

In the population of 113 subjects for this study, those with the minor C allele of rs4251961 polymorphism in IL1RN were more likely to be Caucasian, hypertensive, and to be afflicted with coronary heart disease. Higher plasma IL-1ra was associated with an increased risk of infection (other than pneumonia), and the minor C allele of rs4251961 was independently associated with a decreased risk of infection (other than pneumonia). Initial plasma IL-1ra was not predictive of long-term outcome, but patients with the minor C allele of rs4251961 were more likely to experience good (modified Rankin Score <2) long-term outcome.

Conclusions

These data indicate that IL-1ra and IL1RN may influence the risk of infection after stroke, but this influence seems limited to infections other than pneumonia. Further studies are needed to better understand the complexities of immune regulation on infection and outcome after stroke.     

Endovascular Exclusion of a Postendarterectomy Carotid Pseudoaneurysm

Carotid pseudoaneurysms are rare occurrences. They often result from trauma, but can also present following carotid endarterectomy. Treating such pseudoaneurysms can be difficult due to previous surgery and limited access to the high internal carotid artery. A case involving a postendarterectomy carotid pseudoaneurysm treated via a femoral approach with a covered stent using endoluminal techniques is presented.     

Macronutrient intake in preschoolers with cystic fibrosis and the relationship between macronutrients and growth

Background

Adequate nutrition is essential for growth in children with cystic fibrosis (CF). The new CF Foundation Clinical Practice Guidelines bring attention to monitoring macronutrient intake as well as total energy.

The effectiveness of a pain and anxiety protocol to treat the acute pediatric burn patient

This retrospective review of 286 acute pediatric burn survivors treated in 2001 evaluated the effectiveness of a pharmacotherapeutic protocol for pain, anxiety, and itching. Background pain, procedural pain, exercise pain, anxiety, incidence of acute stress disorder (ASD), and itch were measured with standardized instruments. When this review was compared to similar reviews done in 1993-1994 and 1998, a steady trend toward using more potent pain medications in this patient population is evident. While the use of acetaminophen alone decreased from 50.6% of patients in 1993-1994 and 26.3% in 1998 to 7.3% in 2001, the use of opiates increased from 44.8% in 1993-1994 and 66.9% in 1998 to 81.3% of patients in 2001. Likewise, the use of benzodiazepines for anxiety has increased from 59.8% in 1998 to 77.5% of patients in 2001. During that same period the incidence of ASD decreased from 12.1% in 1993-1994 to 8.7% of patients in 2001. For effective pain and anxiety management, the average administered dose of lorazepam and morphine also increased, providing impetus to revise the pharmacotherapeutic pain protocol. Having a standard pain protocol furnishes a framework for periodic review and facilitates updating of pain and anxiety treatment practices.