Predictors of hippocampal tauopathy in people with and at risk for human immunodeficiency virus infection.

Combination antiretroviral therapy (cART) has extended lifespans of people living with HIV (PWH), increasing both the risk for age-related neuropathologies and the importance of distinguishing effects of HIV and its comorbidities from neurodegenerative disorders. The accumulation of hyperphosphorylated tau (p-tau) in hippocampus is a common degenerative change, with specific patterns of hippocampal subfield vulnerability observed in different disease contexts. Currently, associations between chronic HIV, its comorbidities, and p-tau burden and distribution in the hippocampus are unexplored. We used immunohistochemistry with antibody AT8 to analyze hippocampal p-tau in brain tissues of PWH (n = 71) and HIV negative controls (n = 25), for whom comprehensive clinical data were available. Using a morphology-based neuroanatomical segmentation protocol, we annotated digital slide images to measure percentage p-tau areas in the hippocampus and its subfields. Factors predicting p-tau burden and distribution were identified in univariate analyses, and those with significance at p ≤ 0.100 were advanced to multivariable regression. The patient sample had a mean age of 61.5 years. Age predicted overall hippocampal p-tau burden. Subfield p-tau predictors were for Cornu Ammonis (CA)1, age; for CA2 and subiculum, seizure history; for CA3, seizure history and head trauma; and for CA4/dentate, history of hepatitis C virus (HCV) infection. In this autopsy sample, hippocampal p-tau burden and distribution were not predicted by HIV, viral load, or immunologic status, with viral effects limited to associations between HCV and CA4/dentate vulnerability. Hippocampal p-tau pathologies in cART-era PWH appear to reflect age and comorbidities, but not direct effects of HIV infection.

Frontal lobe microglia, neurodegenerative protein accumulation, and cognitive function in people with HIV.

Microglia are implicated in Alzheimer's Disease (AD) pathogenesis. In a middle-aged cohort enriched for neuroinflammation, we asked whether microgliosis was related to neocortical amyloid beta (A[Formula: see text]) deposition and neuronal phosphorylated tau (p-tau), and whether microgliosis predicted cognition. Frontal lobe tissue from 191 individuals autopsied with detectable (HIV-D) and undetectable (HIV-U) HIV infection, and 63 age-matched controls were examined. Immunohistochemistry (IHC) was used to evaluate A[Formula: see text] plaques and neuronal p-tau, and quantitate microgliosis with markers Iba1, CD163, and CD68 in large regions of cortex. Glia in the A[Formula: see text] plaque microenvironment were quantitated by immunofluorescence (IF). The relationship of microgliosis to cognition was evaluated. No relationship between A[Formula: see text] or p-tau accumulation and overall severity of microgliosis was discerned. Individuals with uncontrolled HIV had the greatest microgliosis, but fewer A[Formula: see text] plaques; they also had higher prevalence of APOE [Formula: see text]4 alleles, but died earlier than other groups. HIV group status was the only variable predicting microgliosis over large frontal regions. In contrast, in the A[Formula: see text] plaque microenvironment, APOE [Formula: see text]4 status and sex were dominant predictors of glial infiltrates, with smaller contributions of HIV status. Cognition correlated with large-scale microgliosis in HIV-D, but not HIV-U, individuals. In this autopsy cohort, over large regions of cortex, HIV status predicts microgliosis, whereas in the A[Formula: see text] plaque microenvironment, traditional risk factors of AD (APOE [Formula: see text]4 and sex) are stronger determinants. While microgliosis does not predict neurodegenerative protein deposition, it does predict cognition in HIV-D. Increased neuroinflammation does not initiate amyloid deposition in a younger group with enhanced genetic risk. However, once A[Formula: see text] deposits are established, APOE [Formula: see text]4 predicts increased plaque-associated inflammation.

HIV disease duration, but not active brain infection, predicts cortical amyloid beta deposition.

OBJECTIVE: Abnormal deposition of the antimicrobial peptide amyloid beta (Aß) is a characteristic of Alzheimer's disease. The objective of this study was to elucidate risk factors for brain Aß in a cohort enriched for HIV and other neurotropic pathogens. DESIGN: Cross-sectional cohort study. METHODS: We examined autopsy brains of 257 donors with a mean age of 52.8 years; 62% were men; and 194 were HIV+ and 63 HIV-. Hyperphosphorylated tau (p-tau) and Aß were identified in frontal and temporal regions by immunohistochemistry. APOE genotyping was performed. Clinical and neuropathological predictors for Aß were identified in univariate analyses, and then tested in multivariate regressions. RESULTS: Cortical Aß was identified in 32% of the sample, and active brain infection in 27%. Increased odds of Aß were seen with increasing age and having an APOE ε4 allele; for the overall sample, HIV+ status was protective and brain infection was not a predictor. Within the HIV+ population, predictors for Aß were duration of HIV disease and APOE alleles, but not age. When HIV disease duration and other HIV parameters were introduced into models for the entire sample, HIV disease duration was equivalent to age as a predictor of Aß. CONCLUSION: We hypothesize that dual aspects of immune suppression and stimulation in HIV, and beneficial survivor effects in older HIV+ individuals, account for HIV+ status decreasing, and HIV duration increasing, odds of Aß. Importantly, with HIV, disease duration replaces age as an independent risk for Aß, suggesting HIV-associated accelerated brain senescence.

Alzheimer's disease neuropathology may not predict functional impairment in HIV: a report of two individuals.

With aging of HIV populations, there is concern that Alzheimer's disease (AD) may become prevalent and difficult to distinguish from HIV-associated neurocognitive disorders. To date, there are no reports documenting histologically verified Alzheimer's neuropathology in individuals with HIV and dementia. Herein, we report two antiretroviral-treated, virally suppressed, HIV-infected individuals autopsied by the Manhattan HIV Brain Bank. Subject A presented to study at 52 years, already dependent in instrumental activities of daily living (ADLs), with severe cognitive impairment inclusive of learning and memory dysfunction. Her history was significant for educational disability and head trauma. She had rapid cognitive decline and, by death at age 59 years, was bed-bound, incontinent, and non-communicative. At autopsy, she exhibited severe AD neuropathologic change (NIA-AA score A3B3C3) and age-related tau astrogliopathy (ARTAG). She was homozygous for APOE ε3/ε3. No HIV DNA was detected in frontal lobe by nested polymerase chain reaction. Subject B was a community dwelling 81-year-old woman who experienced sudden death by pulmonary embolus. Prior to death, she was fully functional, living independently, and managing all ADLs. At autopsy, she displayed moderate amyloid and severe tau AD neuropathologic changes (A2B3C2), ARTAG, and cerebral congophilic angiopathy. She was an APOE ε3/ε4 heterozygote, and HIV DNA, but not RNA, was detected in frontal lobe, despite 20 years of therapy-induced viral suppression. We conclude that in the setting of HIV, AD neuropathology may occur with or without symptomatic cognitive dysfunction; as with seronegative individuals, there are likely to be complex factors in the generation of clinically relevant impairments.

Sprifermin treatment enhances cartilage integration in an in vitro repair model.

Cartilage integration remains a clinical challenge for treatment of focal articular defects. Cartilage exhibits limited healing capacity that declines with tissue maturation. Many approaches have been investigated for their ability to stimulate healing of mature cartilage or integration of repair tissue or tissue-engineered constructs with native cartilage. Growth factors present in immature tissue may enhance chondrogenesis and promote integrative repair of cartilage defects. In this study, we assessed the role of one such factor, fibroblast growth factor 18 (FGF18). Studies using FGF18 have shown a variety of positive effects on cartilage, including stimulation of chondrocyte proliferation, matrix biosynthesis, and suppression of proteinase activity. To explore the role of FGF18 on cartilage defect repair, we hypothesized that treatment with recombinant human FGF18 (sprifermin) would increase matrix synthesis in a defect model, thus improving integration strength. To test this hypothesis, 6 mm cartilage cylinders were harvested from juvenile bovine knees. A central 3 mm defect was created in each explant, and this core was removed and replaced. Resulting constructs were cultured in control or sprifermin-containing medium (weekly 24-h exposure of 100 ng/ml sprifermin) for 4 weeks. Mechanical testing, biochemical analysis, micro-CT, scanning electron microscopy, and histology were used to assess matrix production, adhesive strength, and structural properties of the cartilage-cartilage interface. Results showed greater adhesive strength, increased collagen content, and larger contact areas between core and annular cartilage in the sprifermin-treated group. These findings present a novel treatment for cartilage injuries that have potential to enhance defect healing and lateral cartilage-cartilage integration. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2648-2656, 2018.

Biphasic Finite Element Modeling Reconciles Mechanical Properties of Tissue-Engineered Cartilage Constructs Across Testing Platforms.

Cartilage tissue engineering is emerging as a promising treatment for osteoarthritis, and the field has progressed toward utilizing large animal models for proof of concept and preclinical studies. Mechanical testing of the regenerative tissue is an essential outcome for functional evaluation. However, testing modalities and constitutive frameworks used to evaluate in vitro grown samples differ substantially from those used to evaluate in vivo derived samples. To address this, we developed finite element (FE) models (using FEBio) of unconfined compression and indentation testing, modalities commonly used for such samples. We determined the model sensitivity to tissue radius and subchondral bone modulus, as well as its ability to estimate material parameters using the built-in parameter optimization tool in FEBio. We then sequentially tested agarose gels of 4%, 6%, 8%, and 10% weight/weight using a custom indentation platform, followed by unconfined compression. Similarly, we evaluated the ability of the model to generate material parameters for living constructs by evaluating engineered cartilage. Juvenile bovine mesenchymal stem cells were seeded (2 × 107 cells/mL) in 1% weight/volume hyaluronic acid hydrogels and cultured in a chondrogenic medium for 3, 6, and 9 weeks. Samples were planed and tested sequentially in indentation and unconfined compression. The model successfully completed parameter optimization routines for each testing modality for both acellular and cell-based constructs. Traditional outcome measures and the FE-derived outcomes showed significant changes in material properties during the maturation of engineered cartilage tissue, capturing dynamic changes in functional tissue mechanics. These outcomes were significantly correlated with one another, establishing this FE modeling approach as a singular method for the evaluation of functional engineered and native tissue regeneration, both in vitro and in vivo.

Fibrous Scaffolds with Varied Fiber Chemistry and Growth Factor Delivery Promote Repair in a Porcine Cartilage Defect Model.

Current clinically approved methods for cartilage repair are generally based on either endogenous cell recruitment (e.g., microfracture) or chondrocyte delivery (e.g., autologous chondrocyte implantation). However, both methods culminate in repair tissue with inferior mechanical properties and the addition of biomaterials to these clinical interventions may improve their efficacy. To this end, the objective of this study was to investigate the ability of multipolymer acellular fibrous scaffolds to improve cartilage repair when combined with microfracture in a large animal (i.e., minipig) model. Composite scaffolds were formulated from a combination of hyaluronic acid (HA) fibers and poly(ɛ-caprolactone) (PCL) fibers, either with or without transforming growth factor-ß3 (TGFß3). After 12 weeks in vivo, material choice and TGFß3 delivery had a significant impact on outcomes; specifically, PCL scaffolds without TGFß3 had inferior gross appearance and reduced mechanical properties, whereas HA scaffolds that released TGFß3 resulted in improved histological scores and increased type 2 collagen content. Importantly, analysis of the overall dataset revealed that histology, but not gross appearance, was a better predictor of mechanical properties. This study highlights the importance of scaffold properties on in vivo cartilage repair as well as the need for numerous quantitative outcome measures to fully evaluate treatment methods.

Cartilage repair and subchondral bone remodeling in response to focal lesions in a mini-pig model: implications for tissue engineering.

OBJECTIVE: Preclinical large animal models are essential for evaluating new tissue engineering (TE) technologies and refining surgical approaches for cartilage repair. Some preclinical animal studies, including the commonly used minipig model, have noted marked remodeling of the subchondral bone. However, the mechanisms underlying this response have not been well characterized. Thus, our objective was to compare in-vivo outcomes of chondral defects with varied injury depths and treatments. DESIGN: Trochlear chondral defects were created in 11 Yucatan minipigs (6 months old). Groups included an untreated partial-thickness defect (PTD), an untreated full-thickness defect (FTD), and FTDs treated with microfracture, autologous cartilage transfer (FTD-ACT), or an acellular hyaluronic acid hydrogel. Six weeks after surgery, micro-computed tomography (µCT) was used to quantitatively assess defect fill and subchondral bone remodeling. The quality of cartilage repair was assessed using the ICRS-II histological scoring system and immunohistochemistry for type II collagen. A finite element model (FEM) was developed to assess load transmission. RESULTS: Using µCT, substantial bone remodeling was observed for all FTDs, but not for the PTD group. The best overall histological scores and greatest type II collagen staining was found for the FTD-ACT and PTD groups. The FEM confirmed that only the FTD-ACT group could initially restore appropriate transfer of compressive loads to the underlying bone. CONCLUSIONS: The bony remodeling observed in this model system appears to be a biological phenomena and not a result of altered mechanical loading, with the depth of the focal chondral defect (partial vs. full thickness) dictating the bony remodeling response. The type of cartilage injury should be carefully controlled in studies utilizing this model to evaluate TE approaches for cartilage repair.

Effect of ulnar ostectomy on intra-articular pressure mapping and contact mechanics of the congruent and incongruent canine elbow ex vivo.

OBJECTIVE: To determine (1) the effect of elbow incongruity on contact mechanics and (2) the effect of treatment of this incongruity with 1 of 2 ulnar ostectomies in the canine elbow. STUDY DESIGN: Ex vivo biomechanical study. SAMPLE POPULATION: Unpaired cadaveric canine forelimbs (n = 17). METHODS: In a servohydraulic testing frame, thin-film pressure sensors were placed into the lateral and medial compartments of the elbow. Specimens were tested in 135° of elbow joint flexion at 200 N of cyclic axial force, followed by a 20 seconds hold. Intra-articular contact area (CA), mean contact pressure (mCP) and peak contact pressure (pCP) were measured in each compartment. After radial shortening, testing was repeated and limbs randomized into proximal ulnar ostectomy with IM pin (PUO) or sequential distal ulnar ostectomy (DUO), interosseous ligament release (DUO-L), and ulnar attachment of the abductor pollicis longus muscle and interosseous membrane release (DUO-ML). Paired t-tests were used to compare each treatment to baseline values. Differences between treatment groups were evaluated with a mixed model with random effect to adjust for the clustering of limbs within dog. P < .05 was considered significant. RESULTS: Radial shortening resulted in shift of mCP and pCP from the lateral to the medial compartment. The PUO group resulted in normalization of medial compartment mCP and decrease of pCP, whereas in the DUO group return to baseline was achieved only after DUO-ML. CONCLUSION: PUO is effective in unloading medial compartment pCP in an incongruent joint.

A high throughput mechanical screening device for cartilage tissue engineering.

Articular cartilage enables efficient and near-frictionless load transmission, but suffers from poor inherent healing capacity. As such, cartilage tissue engineering strategies have focused on mimicking both compositional and mechanical properties of native tissue in order to provide effective repair materials for the treatment of damaged or degenerated joint surfaces. However, given the large number design parameters available (e.g. cell sources, scaffold designs, and growth factors), it is difficult to conduct combinatorial experiments of engineered cartilage. This is particularly exacerbated when mechanical properties are a primary outcome, given the long time required for testing of individual samples. High throughput screening is utilized widely in the pharmaceutical industry to rapidly and cost-effectively assess the effects of thousands of compounds for therapeutic discovery. Here we adapted this approach to develop a high throughput mechanical screening (HTMS) system capable of measuring the mechanical properties of up to 48 materials simultaneously. The HTMS device was validated by testing various biomaterials and engineered cartilage constructs and by comparing the HTMS results to those derived from conventional single sample compression tests. Further evaluation showed that the HTMS system was capable of distinguishing and identifying 'hits', or factors that influence the degree of tissue maturation. Future iterations of this device will focus on reducing data variability, increasing force sensitivity and range, as well as scaling-up to even larger (96-well) formats. This HTMS device provides a novel tool for cartilage tissue engineering, freeing experimental design from the limitations of mechanical testing throughput.