No abstract

The role of human bone marrow (BM) CD8+ T cells in the immune response to viral Ags is poorly defined. We report here the identification and characterization of a functionally enhanced effector memory CD8+ T cell population (TEM) in the BM of patients undergoing total joint replacement for osteoarthritis. These BM-derived TEM differ strikingly from correlate cells in peripheral blood (PB), expressing elevated levels of CD27, HLA-DR, CD38, CD69, and unique patterns of chemokine receptors. Interestingly, while BM TEM have low levels of resting perforin and granzyme B, these molecules evidence profound up-regulation in response to TCR stimulation resulting in enhanced cytotoxic potential. Moreover, compared with the TEM subset in PB, BM CD8+ TEM cells demonstrate a more vigorous recall response to pooled viral Ags. Our results reveal that human BM serves as a repository for viral Ag-specific TEM with great therapeutic potential in vaccine development.

Objectives/Hypothesis: Design and test a novel biomaterial for injection laryngoplasty aimed to increase the duration of effectiveness of micronized acellular dermis.Study Design: Animal model. Methods: Injection laryngoplasty was performed in three groups (n 5 5) of New Zealand White rabbits. Acellular dermis was either used alone as a control (group 1), was combined with undifferentiated stem cells (group 2), or with predifferentiated chondrocytic cells (group 3). Groups 2 and 3 were supplemented with growth factors. Animals were sacrificed 4 and 12 weeks after laryngoplasty and histologic analysis was completed. The major outcome measure was volume of tissue remaining.Results: After 4 weeks, the mean volume of tissue remaining was 341 6 89 mm 3 , 295 6 102 mm 3 , and 133 6 15 mm 3 , for groups 1 to 3, respectively. At the 12-week time point, volumes were 62 6 62 mm 3 , 235 6 35 mm 3 , and 107 6 99 mm 3 . After 12 weeks, there was a significantly higher volume in group 2 compared to group 1 or 3 (P 5 .01, P 5 .04). Volumes between week 4 and week 12 were significantly lower in group 1 (P 5 .02), but not significantly different for groups 2 and 3 (P 5 .38, P 5 .74). Histologic evaluation revealed a robust lymphocytic infiltration in all cases as well as morphologic and immunophenotypic features suggestive of chondrogenic differentiation in a single animal.Conclusions: Micronized acellular dermis combined with stem cells and growth factors showed significantly less resorption 12 weeks after injection laryngoplasty compared to micronized acellular dermis alone. Groups using novel tissue-engineered biomaterial showed a lower resorption rate over time compared with acellular dermis alone.

NA Laryngoscope, 127:E166-E169, 2017.

Hypothesis B7-H1 is expressed in vestibular schwannomas. Background Little is known about how benign human vestibular schwannomas interact with antibody-mediated or cell-mediated immunity. We report on the aberrant expression of a novel T-cell coregulatory molecule, B7 homolog 1 (B7-H1), in vestibular schwannomas and discuss the implications of B7-H1 expression and tumor aggressiveness and a potential regulator of B7-H1 expression. Methods Immunohistochemical staining for B7-H1, CD8+, CD3+, and CD4+ lymphocytes were performed on 48 fresh-frozen vestibular schwannoma tissue specimens. A clinical review of patient presenting symptoms and tumor characteristics was performed. Real-time polymerase chain reaction was used to determine if there was differential expression of B7-H1 messenger RNA and microRNA-513, a known regulator of B7-H1, in several strongly positive and negative B7-H1 vestibular schwannomas. Results Nine (19%) of 48 tumors were negative, 23 (48%) tumors were 1+ mildly positive (<20% section area), and 16 (33%) stained 2+ strongly positive (≥20% section area) for B7-H1. The average number of CD8+ cells per high-power field was 2.1 for positive-staining tumors and 1.0 for negative tumors (p = 0.16). Failure of tumor control with stereotactic radiation (p = 0.029) was significantly greater in the strongly positive B7-H1 tumors. Real-time polymerase chain reaction did not show significant differential expression of microRNA-513 (p = 0.62) or B7-H1 messenger RNA (p = 0.35) between the tumors showing strong and negative immunohistochemical staining for B7-H1 protein. Conclusion Vestibular schwannoma tumors express B7-H1, which has been associated with immune tolerance and adverse disease characteristics in several malignancies. Growing tumors that were surgically removed after failed stereotactic radiation therapy were significantly more likely to strongly express B7-H1 protein, which lends some credibility to the hypothesis that immuno-evasion may play some role in their continued growth. Although clinical trends were seen, greater statistical power is required to evaluate whether B7-H1 expression correlates with more aggressive tumor growth or poorer hearing class. B7-H1 seems to be expressed in equal amounts at the RNA level in all vestibular schwannoma tumors that suggests that differential protein expression is occurring at the posttranscriptional level. However, microRNA-513 does not regulate B7-H1 protein expression in these tumors.

Objectives/Hypothesis: Test a new jellyfish collagen biomaterial aimed to increase duration of injection medialization laryngoplasty (IL) against two products in clinical practice.Study Design: Animal model. Methods: Left recurrent laryngeal nerve sectioning and IL were performed in New Zealand White rabbits (N = 6/group). Group 1 received micronized cross-linked jellyfish collagen (MX-JC) and adipose derived stem cells (ADSCs), Group 2, MX-JC alone, Group 3, cross-linked hyaluronic acid (X-HA), and Group 4, micronized acellular dermis (MACD). Animals were sacrificed at 4 and 12 weeks. Major outcomes were MRI tissue volumes and histopathology.Results: After 100 μL IL MRI volumes (means AE STD) at 4 and 12 weeks were:

Objective Human adipose-derived mesenchymal stem cells (ADSCs) were used to rehabilitate bone damaged by osteoradionecrosis (ORN) in an established animal model. Study Design Prospective animal study. Setting Academic department laboratory. Subjects and Methods After institutional review board and Institutional Animal Care and Use Committee approval, 24 athymic nude rats were divided into 5 groups: 4 groups irradiated (20 Gy) by brachytherapy catheter placed at the left hemimandible and 1 mock irradiation control (n = 4). For all groups, ORN was initiated by extraction of the central molar 1 week later. After 28 days, animals (n = 5/group) received injection at the extraction site with saline (SAL), ADSCs, platelet-rich plasma and collagen (PRP/COL), or ADSCs + PRP/COL. Rats were sacrificed 28 days later and their mandibles harvested for histopathology analysis (osteoblasts, osteoclasts, and fibrosis) and bone volume measurement using 3-dimensional micro-computed tomography. Results All but 1 rat survived the experiment period (23/24). Radiographic and histological analysis revealed 60% bone loss in the SAL group compared with the nonirradiated control. Injection of ADSCs increased jaw region bone volume by up to 36% ( P < .01). All experimental groups (ADSC, PRP/COL, and ADSC + PRP/COL) showed dramatically decreased osteoclast counts ( P < .001) while injection of PRP/COL with or without ADSCs increased osteoblasts. Increased fibrosis was observed after ADSC injection ( P < .05). Conclusion The application of human ADSCs to an induced mandibular osteoradionecrosis model in athymic rats results in increased deposition or preservation of bone, demonstrated both histologically and radiographically. This offers an encouraging possible treatment option for translational research in this difficult disease.

IMPORTANCE Nasal reconstruction in patients who are missing a significant amount of structural nasal support remains a difficult challenge. One challenge is the deficiency of cartilage left within the nose as a consequence of rhinectomy or a midline destructive disease. Historically, the standard donor source for large quantities of native cartilage has been costal cartilage.OBJECTIVE To enable the development of protocols for new mesenchymal stem cell technologies as alternative procedures with reduced donor site morbidity, risk of infection and extrusion. DESIGN, SETTING, AND MATERIALSWe examined 6 popular scaffold materials in current practice in terms of their biodegradability in tissue culture, effect on adipose-derived mesenchymal stem cell growth, and chondrogenic fate commitment. Various biomaterials of matching size, porosity, and fiber alignment were synthesized by electrospinning and overlaid with rabbit adipose-derived mesenchymal cells in media supplemented or not with chondrogenic factors. Experiments were performed in vitro using as end points biomarkers for cell growth and chondrogenic differentiation. Polydioxanone (PDO), poly-3hydroxybutyrate-co-3-hydroxyvalerate (PHBV), PHBV-polycaprolactone, poly(L-lactide-cocaprolactone), poly(lactic-co-glycolic acid), and polystyrene scaffolds of 60% to 70% porosity and random fiber alignment were coated with poly(L)-lysine/laminin to promote cell adhesion and incubated for 28 days with 2.5 to 3.5 × 10 5 rabbit adipose mesenchymal cells. MAIN OUTCOMES AND MEASURESCell growth was measured by fluorometric DNA quantitation and chondrogenic differentiation of stem cells by spectrophotometric sulfated glycosaminoglycan (sGAG) assay. Microscopic visualization of cell growth and matrix deposition on formalin-fixed, paraffin-embedded tissue sections was performed, respectively, with nuclear fast red and Alcian blue. RESULTSOf 6 scaffold materials tested using rabbit apidose mesenchymal cells, uncoated scaffolds promoted limited cell adhesion but coating with poly(L)-lysine/laminin enabled efficient cell saturation of scaffold surfaces, albeit with limited involvement of scaffold interiors. Similar growth rates were observed under these conditions, based on DNA content analysis. However, PDO and PHBV/PCL scaffolds supported chondrogenic fate commitment better than other materials, based on soluble sGAG analysis and microscopic observation of chondrogenic matrix deposition. The mean (SD) sGAG scaffold values expressed as fold increase over control were PDO, 2.26 (0.88), PHBV/PCL, 2.09 (0.83), PLCL, 1.36 (0.39), PLGA, 1.34 (0.77), PHBV, 1.07 (0.31), and PS, 0.38 (0.14). CONCLUSIONS AND RELEVANCEThese results establish materials, reagents, and protocols for tissue engineering for nasal reconstruction using single-layer, chondrogenically differentiated, adipose-derived mesenchymal stem cells. Stackable, scaffold-supported, multisheet bioengineered tissue may be generated using these protocols.LEVEL OF EVIDENCE NA.