A morphological and molecular approach to investigating infectious disease in early medieval Iberia: The necropolis of La Olmeda (Palencia, Spain).

OBJECTIVE: Here we investigate infectious diseases that potentially contribute to osteological lesions in individuals from the early medieval necropolis of La Olmeda (6th-11th c. CE) in North Iberia. MATERIALS AND METHODS: We studied a minimum number of 268 individuals (33 adult females; 38 adult males, 77 unknown/indeterminate sex; and 120 non-adults), including articulated and commingled remains. Individuals with differential diagnoses suggesting chronic systemic infectious diseases were sampled and bioinformatically screened for ancient pathogen DNA. RESULTS: Five non-adults (and no adults) presented skeletal evidence of chronic systemic infectious disease (1.87% of the population; 4.67% of non-adults). The preferred diagnoses for these individuals included tuberculosis, brucellosis, and malaria. Ancient DNA fragments assigned to the malaria-causing pathogen, Plasmodium spp., were identified in three of the five individuals. Observed pathology includes lesions generally consistent with malaria; however, additional lesions in two of the individuals may represent hitherto unknown variation in the skeletal manifestation of this disease or co-infection with tuberculosis or brucellosis. Additionally, spondylolysis was observed in one individual with skeletal lesions suggestive of infectious disease. CONCLUSIONS: This study sheds light on the pathological landscape in Iberia during a time of great social, demographic, and environmental change. Genetic evidence challenges the hypothesis that malaria was absent from early medieval Iberia and demonstrates the value of combining osteological and archaeogenetic methods. Additionally, all of the preferred infectious diagnoses for the individuals included in this study (malaria, tuberculosis, and brucellosis) could have contributed to the febrile cases described in historical sources from this time.

Multinucleated giant cells within the in vivo implantation bed of a collagen-based biomaterial determine its degradation pattern.

OBJECTIVES: The aim of the present study was to characterize the cellular reaction to a xenogeneic resorbable collagen membrane of porcine origin using a subcutaneous implantation model in Wistar rats over 30 days. MATERIALS AND METHODS: Ex vivo, liquid platelet-rich fibrin (PRF), a leukocyte and platelet-rich cell suspension, was used to evaluate the blood cell membrane interaction. The material was implanted subcutaneously in rats. Sham-operated rats without biomaterial displayed physiological wound healing (control group). Histological, immunohistological, and histomorphometric analyses were focused on the inflammatory pattern, vascularization rate, and degradation pattern. RESULTS: The membrane induced a large number of mononuclear cells over the observation period, including lymphocytes, macrophages, and fibroblasts. After 15 days, multinucleated giant cells (MNGCs) were observed on the biomaterial surface. Their number increased significantly, and they proceeded to the center of the biomaterial on day 30. These cells highly expressed CD-68, calcitonin receptor, and MMP-9, but not TRAP or integrin-ß3. Thus, the membrane lost its integrity and underwent disintegration as a consequence of the induction of MNGCs. The significant increase in MNGC number correlated with a high rate of vascularization, which was significantly higher than the control group. Physiological wound healing in the control group did not induce any MNGCs at any time point. Ex vivo blood cells from liquid-PRF did not penetrate the membrane. CONCLUSION: The present study suggests a potential role for MNGCs in biomaterial degradation and questions whether it is beneficial to accept them in clinically approved biomaterials or focus on biomaterials that induce only mononuclear cells. Thus, further studies are necessary to identify the function of biomaterial-induced MNGCs. CLINICAL RELEVANCE: Understanding the cellular reaction to biomaterials is essential to assess their suitability for specific clinical indications and outline the potential benefit of specific group of biomaterials in the respective clinical indications.

Excellent histological results in terms of articular cartilage regeneration after spheroid-based autologous chondrocyte implantation (ACI).

PURPOSE: Traumatic lesions of articular cartilage represent a crucial risk factor for osteoarthritis. Even if several strategies exist to treat such damages, the optimal solution has not yet been found. A new strategy represents the scaffold-free spheroid-based autologous chondrocyte transplantation. In this method, spheroids of chondrocytes are synthesized after chondrocyte isolation and expansion, followed by the implantation in a second intervention. METHODS: Fine Jamshidi-needle biopsies from five patients (one from each patient, Ø 2 mm) treated with a spheroid-based autologous chondrocyte implantation (ACI) after traumatic lesions of the articular cartilage of the knee were analysed histologically and immunohistologically for collagen II, collagen X and aggrecan expression. The indication for a second look arthroscopy was given by arthrofibrosis or meniscus-lesions, respectively. The time between ACI and second-look arthroscopy ranged between 6 and 16 months. RESULTS: In all patients, the histological examinations revealed an avascular cartilage tissue with a homogenic extracellular matrix. The subchondral bone neither showed bleeding, necrosis nor hypertrophy. A homogenous alcian blue staining indicated high amounts of mucopolysaccharides and glycosaminoglycans. Collagen II staining was highly positive, whereas collagen X staining was negative in every patient, ruling out hypertrophic chondrocyte differentiation. In addition, intense aggrecan staining indicated a strong expression of this extracellular matrix component. CONCLUSION: The present case series represents the first histological and immunohistological analyses of spheroid-based ACI in humans. Spheroid-based ACI revealed excellent histological results regarding the regeneration of hyaline articular cartilage. These results indicate that spheroid based ACI is a promising strategy for treating traumatic lesions of the articular cartilage of the knee.

Characterization of the Cellular Reaction to a Collagen-Based Matrix: An In Vivo Histological and Histomorphometrical Analysis.

The permeability and inflammatory tissue reaction to Mucomaix® matrix (MM), a non- cross-linked collagen-based matrix was evaluated in both ex vivo and in vivo settings. Liquid platelet rich fibrin (PRF), a blood concentrate system, was used to assess its capacity to absorb human proteins and interact with blood cells ex vivo. In the in vivo aspect, 12 Wister rats had MM implanted subcutaneously, whereas another 12 rats (control) were sham-operated without biomaterial implantation. On days 3, 15 and 30, explantation was completed (four rats per time-point) to evaluate the tissue reactions to the matrix. Data collected were statistically analyzed using analysis of variance (ANOVA) and Tukey multiple comparisons tests (GraphPad Prism 8). The matrix absorbed the liquid PRF in the ex vivo study. Day 3 post-implantation revealed mild tissue inflammatory reaction with presence of mononuclear cells in the implantation site and on the biomaterial surface (mostly CD68-positive macrophages). The control group at this stage had more mononuclear cells than the test group. From day 15, multinucleated giant cells (MNGCs) were seen in the implantation site and the outer third of the matrix with marked increase on day 30 and spread to the matrix core. The presence of these CD68-positive MNGCs was associated with significant matrix vascularization. The matrix degraded significantly over the study period, but its core was still visible as of day 30 post-implantation. The high permeability and fast degradation properties of MM were highlighted.

Function and mutual interaction of BiP-, PERK-, and IRE1α-dependent signalling pathways in vascular tumours.

Spontaneously regressing infantile haemangiomas and aggressive angiosarcomas are vascular tumours with excessive angiogenesis. When analysing haemangiomas and angiosarcomas immunohistochemically with respect to their chaperone profiles we found that angiosarcomas have significantly elevated protein levels of binding immunoglobulin protein (BIP) and PERK with concomitant attenuated IRE1α levels, whereas haemangioma tissue exhibits the same pattern as embryonal skin tissue. We show that BiP is essential for the maintenance of VEGFR2 protein, which is expressed in the endothelium of both tumour types. When studying the effects of BiP, the IRE1α/Xbp1 -, and PERK/ATF4-signalling pathways on the migration and tube-forming potential of endothelial cells, we show that downregulation of BiP, as well as inhibition of the kinase activity of IRE1α, inhibit in vitro angiogenesis. Downregulation of PERK (PKR-like kinase; PKR = protein kinase R) levels promotes Xbp1 splicing in endoplasmic reticulum (ER)-stressed cells, indicating that in angiosarcoma the elevated PERK levels might result in high levels of unspliced Xbp1, which have been reported to promote cell proliferation and increase tumour malignancy. The data presented in this study revealed that in addition to BiP or PERK, the kinase domains of IRE1α and Xbp1 could be potential targets for the development of novel therapeutic approaches for treating angiosarcomas and to control tumour angiogenesis. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

The Biomaterial-Induced Cellular Reaction Allows a Novel Classification System Regardless of the Biomaterials Origin.

Several different biomaterials are being introduced for clinical applications. However, no current material-specific systematic studies define parameters for evaluating these materials. The aim of this retrospective animal study is to classify biomaterials according to the in vivo induced cellular reaction and outline the clinical consequence of the biomaterial-specific cellular reaction for the regeneration process. A retrospective histologic analysis was performed for 13 polymeric biomaterials and 19 bone substitute materials (BSMs) (of various compositions and origins) that were previously implanted in a standardized subcutaneous model. Semiquantitative analyses were performed at days 3, 15, and 30 after implantation according to a standardized score for the induction of multinucleated giant cells (MNGCs) and vascularization rate. The induced cellular reaction in response to different polymeric materials allowed their classification according to the MNGC score in the following groups: class I induced no MNGCs at any time point, class II induced and maintained a constant number of MNGCs over 30 days, and class III induced MNGCs and provided an increasing number over 30 days. All BSMs induced MNGCs to varying extents. Therefore, the resultant BSM classifications are as follows: class I induced MNGCs with a decreasing number, class II induced and maintained constant MNGCs over 30 days, and class III induced MNGCs with increasing number over 30 days. These observations were mostly related to the biomaterial physicochemical properties and were independent of the biomaterial origin. Consequently, the induction of MNGCs and their increase over 30 days resulted in disintegration of the biomaterial. By contrast, the absence of MNGCs resulted in an integration of the biomaterial within the host tissue. This novel classification provides clinicians a tool to assess the capacity and suitability of biomaterials in the intended clinical indication for bone and soft tissue implantations.

Modification of collagen-based sponges can induce an upshift of the early inflammatory response and a chronic inflammatory reaction led by M1 macrophages: an in vivo study.

BACKGROUND: The present study evaluated the cellular tissue reaction of two equine-derived collagen hemostatic sponges (E-CHS), which differed in thickness after pressing, over 30 days in vivo. The inflammatory response during physiological wound healing in sham-operated animals was used as control group. MATERIAL AND METHODS: First, the E-CHS was pressed by applying constant pressure (6.47 ± 0.85 N) for 2 min using a sterile stainless-steel cylinder until the material was uniformly flattened. Consequently, the original (E-CHS), the pressed (P-E-CHS), as well as the control group (CG; sham operation) were studied independently. The 3 groups were evaluated in vivo after subcutaneous implantation in Wistar rats during 3, 15, and 30 days. Histochemical and immunohistochemical methods provided observations of biomaterial degradation rate, cellular inflammatory response, and vascularization pattern. A derivative of human blood known as platelet-rich fibrin (PRF) was used as an ex vivo model to simulate the initial biomaterial-cell interaction. Segments of E-CHS and P-E-CHS were cultivated for 3 and 6 days with PRF, and the release of pro-inflammatory proteins was measured using ELISA. PRF cultivated alone was used as a control group. RESULTS: At day 3, the CG induced a statistically significant higher presence of monocytes/macrophages (CD68+), pro-inflammatory macrophages (M1; CCR7+), and pro-wound healing macrophages (M2; CD206+) compared to E-CHS and P-E-CHS. At the same time point, P-E-CHS induced a statistically significant higher presence of CD68+ cells compared to E-CHS. After 15 days, E-CHS was invaded by cells and vessels and showed a faster disintegration rate compared to P-E-CHS. On the contrary, cells and vessels were located only in the outer region of P-E-CHS and the biomaterial did not lose its structure and accordingly did not undergo disintegration. The experimental groups induced similar inflammatory reaction primarily with positive pro-inflammatory CD68+/CCR7+ macrophages and a low presence of multinucleated giant cells (MNGCs). At this time point, significantly lower CD68+/CCR7+ macrophages and no MNGCs were detected within the CG when compared to the experimental groups (P < 0.05). After 30 days, E-CHS and P-E-CHS were fully degraded. All groups showed similar inflammatory reaction shifted to a higher presence CD206+ macrophages. A low number of CCR7+ MNGCs were still observable in the implantation bed of both experimental groups. In the ex vivo model, the cells and fibrin from PRF penetrated E-CHS. However, in the case of P-E-CHS, the cells and fibrin stayed on the surface and did not penetrate towards materials central regions. The cultivation of P-E-CHS with PRF induced a statically significant higher release of pro-inflammatory proteins compared to the CG and E-CHS after 3 days. CONCLUSION: Altering the original presentation of a hemostatic sponge biomaterial by pressing modified the initial biomaterial-cell interaction, delayed the early biomaterial's degradation rate, and altered the vascularization pattern. A pressed biomaterial seems to induce a higher inflammatory reaction at early time points. However, altering the biomaterial did not modify the polarization pattern of macrophages compared to physiologic wound healing. The ex vivo model using PRF was shown to be an effective model to simulate the initial biomaterial-cell interaction in vivo. CLINICAL RELEVANCE: A pressed hemostatic sponge could be applied for guided tissue regeneration and guided bone regeneration. In that sense, within the limitations of this study, the results show that the same biomaterial may have two specific clinical indications.

In Vitro Entero-Capillary Barrier Exhibits Altered Inflammatory and Exosomal Communication Pattern after Exposure to Silica Nanoparticles.

The intestinal microvasculature (iMV) plays multiple pathogenic roles during chronic inflammatory bowel disease (IBD). The iMV acts as a second line of defense and is, among other factors, crucial for the innate immunity in the gut. It is also the therapeutic location in IBD targeting aggravated leukocyte adhesion processes involving ICAM-1 and E-selectin. Specific targeting is stressed via nanoparticulate drug vehicles. Evaluating the iMV in enterocyte barrier models in vitro could shed light on inflammation and barrier-integrity processes during IBD. Therefore, we generated a barrier model by combining the enterocyte cell line Caco-2 with the microvascular endothelial cell line ISO-HAS-1 on opposite sides of a transwell filter-membrane under culture conditions which mimicked the physiological and inflamed conditions of IBD. The IBD model achieved a significant barrier-disruption, demonstrated via transepithelial-electrical resistance (TER), permeability-coefficient (Papp) and increase of sICAM sE-selectin and IL-8. In addition, the impact of a prospective model drug-vehicle (silica nanoparticles, aSNP) on ongoing inflammation was examined. A decrease of sICAM/sE-selectin was observed after aSNP-exposure to the inflamed endothelium. These findings correlated with a decreased secretion of ICAM/E-selectin bearing exosomes/microvesicles, as evaluated via ELISA. Our findings indicate that aSNP treatment of the inflamed endothelium during IBD may hamper exosomal/microvesicular systemic communication.

Multiwell three-dimensional systems enable in vivo screening of immune reactions to biomaterials: a new strategy toward translational biomaterial research.

In vivo experiments are accompanied by ethical issues, including sacrificing a large number of animals as well as large costs. A new in vivo 3D screening system was developed to reduce the number of required animals without compromising the results. The present pilot study examined a multiwell array system in combination with three different collagen-based biomaterials (A, B and C) using subcutaneous implantation for 10 days and histological and histomorphometrical evaluations. The tissue reaction towards the device itself was dominated by mononuclear cells. However, three independent biomaterial-specific tissue reactions were observed in three chambers. The results showed a mononuclear cell-based tissue reaction in one chamber (A) and foreign body reaction by multinucleated giant cells in the other two chambers (B and C). Statistical analysis showed a significantly higher number of multinucleated giant cells in cases B and C than in case A (A vs. B; ***P < 0.001), (A vs. C; P < 0.01). These outcomes were comparable to previously published observations with conventional biomaterial implantation. The present data lead to the conclusion that this 3D screening system could be an alternative tool to enhance the effectiveness of in vivo experiments, thus offering a more economic strategy to screen biomaterial-related cellular reactions, while saving animals, without influencing the final outcome.

Tissue expansion of lung bronchi due to tissue processing for histology - A comparative analysis of paraffin versus frozen sections in a pig model.

AIM: Tissue shrinking due to fixation and processing is well known. However, the degree of shrinking varies significantly with the tissue type as well as the processing method and is not well studied in various tissues. In daily pathological routine workflow, histological specimens from frozen and paraffin sections are performed from the same tissue. In the present study we compared the thickness of bronchus walls obtained from paraffin and frozen sections. METHODS: Pig lungs were frozen in ventilated condition in liquid nitrogen and 36 bronchi were isolated after dissection. Frozen sections of 5 µm thickness were performed and the remaining tissue was fixed and embedded in paraffin after fixation in 4% formalin. Frozen and paraffin sections from the same cutting edge were analysed after haematoxylin and eosin staining by measuring the wall thickness of the bronchi using high power fields of 400-fold magnification. In each bronchus 40 measurements were implemented at different wall positions distributed over the entire wall area. Summed up, in each group 1440 wall measurements were performed in total. Statistical analysis was conducted using the Wilcoxon test and t-test as well as Pearson's correlation coefficient with a significance level at P < 0.05. RESULTS: The bronchial wall thickness was significantly (p < 0.001) smaller in frozen sections (median: 0.50 mm; min: 0.37 mm; max: 0.97 mm) compared to paraffin sections (median: 0.58 mm; min: 0.35 mm; max: 1.06 mm). The median difference between paraffin and frozen sections was 0.05 mm (min: -0.11 mm; max: 0.22 mm). The wall thickness ratio of both groups was as follows: frozen/paraffin section = 0.8609, thus yielding a difference between paraffin and frozen of 13.91%. High correlation was found between wall thickness measurements on paraffin and frozen sections (R = 0.87, p < 0.001). CONCLUSIONS: The bronchus wall thickness in the frozen section was 14% reduced compared to the paraffin section. In routine pathology as well as in scientific studies these results are of relevance, as airway wall thickness represents a relevant marker for pathological interpretation, especially using CT image techniques.

Platelet-rich fibrin secretome induces three dimensional angiogenic activation in vitro.

Different tissue engineering techniques are used to support rapid vascularisation. A novel technique is the use of platelet-rich fibrin (PRF), an autologous source of growth factors. This study was the first to investigate the influence of PRF matrices, isolated following different centrifugation protocols, on human dermal vascular endothelial cells (ECs) in mono-culture and co-culture with human primary fibroblasts (HFs) as an in vitro model for tissue regeneration. Focus was placed on vascular structure formation and growth factor release. HFs and ECs were cultivated with PRF prepared using a high (710 ×g) or low (44 ×g) relative centrifugation force (RCF) over 14 d. Immunofluorescence staining and immunohistochemistry were used to evaluate the microvascular formation. Cell culture supernatants were collected for evaluation of growth factor release. The results showed a PRF-mediated effect on the induction of angiogenesis in ECs. Microvessel-like structure formation was promoted when ECs were combined with low-RCF PRF as compared to high-RCF PRF or control group. The percentage of vascular lumen area was significantly higher in low-RCF PRF, especially at day 7, which coincided with statistically significantly higher growth factor [vascular endothelial factor (VEGF), transforming growth factor ß1 (TGF-ß1) and platelet derived growth factor (PDGF)] concentration measured in low-RCF PRF as compared to high-RCF PRF or control group. In conclusion, reducing the RCF according to the low-speed centrifugation concept (LSCC) resulted in increased growth factor release and angiogenic structure formation with EC mono-culture, suggesting that PRF may be a highly beneficial therapeutic tool for tissue engineering applications.

Evaluation of a pharmacist-led penicillin allergy de-labelling ward round: a novel antimicrobial stewardship intervention.

BACKGROUND: Antibiotic allergy labels (AALs), reported by up to 25% of hospitalized patients, are a significant barrier to appropriate prescribing and a focus of antimicrobial stewardship (AMS) programmes. METHODS: A prospective audit of a pharmacist-led AMS penicillin allergy de-labelling ward round at Austin Health (Melbourne, Australia) was evaluated. Eligible inpatients with a documented penicillin allergy receiving an antibiotic were identified via an electronic medical report and then reviewed by a pharmacist-led AMS team. The audit outcomes evaluated were: (i) AMS post-prescription review recommendations; (ii) direct de-labelling; (iii) inpatient oral rechallenge referral; (iv) skin prick testing/intradermal testing referral; and (v) outpatient antibiotic allergy clinic assessment. RESULTS: Across a 5 month period, 106 patients were identified from a real-time electronic prescribing antibiotic allergy report. The highest rate of penicillin allergy de-labelling was demonstrated in patients who were referred for an inpatient oral rechallenge with 95.2% (n = 21) successfully having their penicillin AAL removed. From the 22 patients with Type A reactions, 63.6% had their penicillin AAL removed. We demonstrated a significant decrease in the prescribing of restricted antibiotics (defined as third- or fourth-generation cephalosporins, fluoroquinolones, glycopeptides, carbapenems, piperacillin/tazobactam, lincosamides, linezolid or daptomycin) in patients reviewed (pre 42.5% versus post 17.9%, P = 0.0002). CONCLUSIONS: A pharmacist-led AMS penicillin allergy de-labelling ward round reduced penicillin AALs and the prescribing of restricted antibiotics. This model could be implemented at other hospitals with existing AMS programmes.

Palmar/plantar approach for radiographic-guided injection of the equine distal interphalangeal joint collateral ligament insertion.

There are limited radiographic-guided injection techniques of the insertion of the distal interphalangeal joint (DIPJ) collateral ligaments. The objective of this study was to develop and evaluate a palmar/plantar radiographic-guided injection of the collateral ligament insertion in cadavers. Fifty limbs were used to develop the technique and 24 additional limbs were used to evaluate accuracy. An 18 G, 9 cm spinal needle was placed in the depression between the palmar digital neurovascular bundle and arch of the ungular cartilage with dorsodistal advancement towards the distal phalanx collateral fossa. Radiographs verified ideal needle location on the proximal border of the distal phalanx at the collateral fossa. Dye was injected. Hoof walls were partially removed and collateral ligaments were dissected with needles in place to determine needle and dye location. Accuracy of needle placement into the insertion of the DIPJ collateral ligament was 41/48 (85 per cent), with lower accuracy of dye within the ligament (34/48; 71 per cent). Dye entered the DIPJ in 2/48 injections, but dye entered periligamentous structures in 22/48 (46 per cent) injections. A palmar/plantar radiographic-guided injection of the insertion of the DIPJ collateral ligament had high accuracy rate with low injection rate of the DIPJ in cadavers.

Reduction of relative centrifugal forces increases growth factor release within solid platelet-rich-fibrin (PRF)-based matrices: a proof of concept of LSCC (low speed centrifugation concept).

Purpose The present study evaluated the platelet distribution pattern and growth factor release (VEGF, TGF-ß1 and EGF) within three PRF (platelet-rich-fibrin) matrices (PRF, A-PRF and A-PRF+) that were prepared using different relative centrifugation forces (RCF) and centrifugation times. Materials and methods immunohistochemistry was conducted to assess the platelet distribution pattern within three PRF matrices. The growth factor release was measured over 10 days using ELISA. Results The VEGF protein content showed the highest release on day 7; A-PRF+ showed a significantly higher rate than A-PRF and PRF. The accumulated release on day 10 was significantly higher in A-PRF+ compared with A-PRF and PRF. TGF-ß1 release in A-PRF and A-PRF+ showed significantly higher values on days 7 and 10 compared with PRF. EGF release revealed a maximum at 24 h in all groups. Toward the end of the study, A-PRF+ demonstrated significantly higher EGF release than PRF. The accumulated growth factor releases of TGF-ß1 and EGF on day 10 were significantly higher in A-PRF+ and A-PRF than in PRF. Moreover, platelets were located homogenously throughout the matrix in the A-PRF and A-PRF+ groups, whereas platelets in PRF were primarily observed within the lower portion. ​Discussion the present results show an increase growthfactor release by decreased RCF. However, further studies must be conducted to examine the extent to which enhancing the amount and the rate of released growth factors influence wound healing and biomaterial-based tissue regeneration. ​Conclusion These outcomes accentuate the fact that with a reduction of RCF according to the previously LSCC (described low speed centrifugation concept), growth factor release can be increased in leukocytes and platelets within the solid PRF matrices.

Biomaterial-induced multinucleated giant cells express proinflammatory signaling molecules: A histological study in humans.

The biomaterials physicochemical characteristics influence their cellular reaction, degradation and regenerative capacities. Macrophages and multinucleated giant cells (MNGCs) are observed in the augmentation area of biomaterials. This study, for the first time, evaluated the polarization pattern of macrophages and MNGCs in response to two different bone substitute materials (synthetic bone substitute material [SBSM] = NanoBone vs. xenogeneic bone substitute material [XBSM] = Bio-Oss) in human bone biopsies compared to non-augmented bone (control). Histomorphometrical analysis of the polarization in proinflammatory (M1) and anti-inflammatory (M2) cells was performed using different immunohistochemical markers: CD-68 = macrophages; CCR-7 and Cox-2 (M1) and CD-206 and CD-163 (M2) and tartrate-resistant acid phosphatase (TRAP). The macrophage polarization pattern in SBSM showed a significantly higher number of M1 cells than did XBSM and non-augmented bone. XBSM induced a significantly higher number of CD-206-positive macrophages than SBSM did. No significant difference was found between XBSM and the non-augmented bone. MNGCs expressed CD-68 and TRAP. In both test-groups, MNGCs showed a high proinflammatory character (CCR-7 and Cox-2-positive) and their number in the SBSM group was significantly higher than that of XBSM. The tissue distribution showed a significantly low percentage of the remaining biomaterial in SBSM compared to XBSM. Within the limitations of this study, these findings show that MNGCs exhibit a rather proinflammatory character and lead to biomaterial degradation, once they are induced in a high number. The premature degradation of bone substitute materials is compensated with a high percentage of connective tissue and not new bone formation. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 780-790, 2019.

A Proof of the Low Speed Centrifugation Concept in Rodents: New Perspectives for In Vivo Research.

IMPACT STATEMENT: This study evaluated for the first time the composition and bioactivity of platelet-rich fibrin (PRF) produced from small animal blood by reducing the initial blood volume needed for the preparation of PRF from 10 to 3 mL. The results showed that different preparation protocols of PRF produced using 3 mL of animal blood exhibit the same composition, properties, and bioactivity as PRF prepared using 10 mL human blood.

Human Co- and Triple-Culture Model of the Alveolar-Capillary Barrier on a Basement Membrane Mimic.

The development of an in vitro model resembling the alveolar-capillary barrier might be a highly beneficial tool to study lung physiology as well as the immune response of the lung to infection or after exposure to nanoparticles. This study is based on an in vitro alveolar barrier developed on a basement membrane mimic, composed of ultrathin nanofiber meshes generated via electrospinning using bioresorbable poly(ɛ-caprolactone). As cellular components, NCI H441, resembling the alveolar epithelial cells, and ISO-HAS-1, an endothelial cell line, were used to perform bipolar coculture experiments for a total cultivation period of 14 days. In addition to immunohistochemical and immunofluorescent studies, transepithelial electrical resistance (TER) and transport capabilities of the in vitro model system were investigated. Alveolar barrier function could be clearly determined for the postulated bipolar coculture system on the basement membrane mimic, since TER increased during the course of bipolar cultivation. Furthermore, to gain first insights into possible lung inflammatory reactions in vitro, this coculture model was further expanded by a human leukemia monocyte cell line (THP-1). This triple-culture system was able to maintain adequately the barrier properties of the bipolar coculture, thus making this in vitro model consisting of epithelial, endothelial, and immune cells on a basement membrane mimic a promising basis for further studies in tissue engineering.

In vivo Implantation of a Bovine-Derived Collagen Membrane Leads to Changes in the Physiological Cellular Pattern of Wound Healing by the Induction of Multinucleated Giant Cells: An Adverse Reaction?

The present study evaluated the tissue response toward a resorbable collagen membrane derived from bovine achilles tendon (test group) in comparison to physiological wound healing (control group). After subcutaneous implantation in Wistar rats over 30 days, histochemical and immunohistochemical methods elucidated the cellular inflammatory response, vascularization pattern, membrane protein and cell absorbance capacity. After 30 days, the test-group induced two different inflammatory patterns. On the membrane surface, multinucleated giant cells (MNGCs) were formed after the accumulation of CD-68-positive cells (macrophages), whereas only mononuclear cells (MNCs) were found within the membrane central region. Peri-implant vascularization was significantly enhanced after the formation of MNGCs. No vessels were found within the central region of the membrane. Physiological wound healing revealed no MNGCs at any time point. These dynamic changes in the cellular reaction and vascularization within the test-group are related typical indications of a foreign body reaction. Due to the membrane-specific porosity, mononuclear cells migrated into the central region, and the membrane maintained its integrity over 30 days by showing no breakdown or disintegration. The ex vivo investigation analyzed the interaction between the membrane and a blood concentrate system, liquid platelet-rich fibrin (liquid PRF), derived from human peripheral blood and consisting of platelets, leukocytes and fibrin. PRF penetrated the membrane after just 15 min. The data question the role of biomaterial-induced MNGCs as a pathological reaction and whether this is acceptable to trigger vascularization or should be considered as an adverse reaction. Therefore, further pre-clinical and clinical studies are needed to identify the types of MNGCs that are induced by clinically approved biomaterials.

Cost-utility analysis of antiviral use under pandemic influenza using a novel approach - linking pharmacology, epidemiology and heath economics.

Simulation models are used widely in pharmacology, epidemiology and health economics (HEs). However, there have been no attempts to incorporate models from these disciplines into a single integrated model. Accordingly, we explored this linkage to evaluate the epidemiological and economic impact of oseltamivir dose optimisation in supporting pandemic influenza planning in the USA. An HE decision analytic model was linked to a pharmacokinetic/pharmacodynamics (PK/PD) - dynamic transmission model simulating the impact of pandemic influenza with low virulence and low transmissibility and, high virulence and high transmissibility. The cost-utility analysis was from the payer and societal perspectives, comparing oseltamivir 75 and 150 mg twice daily (BID) to no treatment over a 1-year time horizon. Model parameters were derived from published studies. Outcomes were measured as cost per quality-adjusted life year (QALY) gained. Sensitivity analyses were performed to examine the integrated model's robustness. Under both pandemic scenarios, compared to no treatment, the use of oseltamivir 75 or 150 mg BID led to a significant reduction of influenza episodes and influenza-related deaths, translating to substantial savings of QALYs. Overall drug costs were offset by the reduction of both direct and indirect costs, making these two interventions cost-saving from both perspectives. The results were sensitive to the proportion of inpatient presentation at the emergency visit and patients' quality of life. Integrating PK/PD-EPI/HE models is achievable. Whilst further refinement of this novel linkage model to more closely mimic the reality is needed, the current study has generated useful insights to support influenza pandemic planning.