FKBPL and SIRT-1 Are Downregulated by Diabetes in Pregnancy Impacting on Angiogenesis and Endothelial Function.

Diabetes in pregnancy is associated with adverse pregnancy outcomes including preterm birth. Although the mechanisms leading to these pregnancy complications are still poorly understood, aberrant angiogenesis and endothelial dysfunction play a key role. FKBPL and SIRT-1 are critical regulators of angiogenesis, however, their roles in pregnancies affected by diabetes have not been examined before in detail. Hence, this study aimed to investigate the role of FKBPL and SIRT-1 in pre-gestational (type 1 diabetes mellitus, T1D) and gestational diabetes mellitus (GDM). Placental protein expression of important angiogenesis proteins, FKBPL, SIRT-1, PlGF and VEGF-R1, was determined from pregnant women with GDM or T1D, and in the first trimester trophoblast cells exposed to high glucose (25 mM) and varying oxygen concentrations [21%, 6.5%, 2.5% (ACH-3Ps)]. Endothelial cell function was assessed in high glucose conditions (30 mM) and following FKBPL overexpression. Placental FKBPL protein expression was downregulated in T1D (FKBPL; p<0.05) whereas PlGF/VEGF-R1 were upregulated (p<0.05); correlations adjusted for gestational age were also significant. In the presence of GDM, only SIRT-1 was significantly downregulated (p<0.05) even when adjusted for gestational age (r=-0.92, p=0.001). Both FKBPL and SIRT-1 protein expression was reduced in ACH-3P cells in high glucose conditions associated with 6.5%/2.5% oxygen concentrations compared to experimental normoxia (21%; p<0.05). FKBPL overexpression in endothelial cells (HUVECs) exacerbated reduction in tubule formation compared to empty vector control, in high glucose conditions (junctions; p<0.01, branches; p<0.05). In conclusion, FKBPL and/or SIRT-1 downregulation in response to diabetic pregnancies may have a key role in the development of vascular dysfunction and associated complications affected by impaired placental angiogenesis.

Role of A Novel Angiogenesis FKBPL-CD44 Pathway in Preeclampsia Risk Stratification and Mesenchymal Stem Cell Treatment.

CONTEXT: Preeclampsia is a leading cardiovascular complication in pregnancy lacking effective diagnostic and treatment strategies. OBJECTIVE: To investigate the diagnostic and therapeutic target potential of the angiogenesis proteins, FK506-binding protein like (FKBPL) and CD44. DESIGN AND INTERVENTION: FKBPL and CD44 plasma concentration or placental expression were determined in women pre- or postdiagnosis of preeclampsia. Trophoblast and endothelial cell function was assessed following mesenchymal stem cell (MSC) treatment and in the context of FKBPL signaling. SETTINGS AND PARTICIPANTS: Human samples prediagnosis (15 and 20 weeks of gestation; n ≥ 57), or postdiagnosis (n = 18 for plasma; n = 4 for placenta) of preeclampsia were used to determine FKBPL and CD44 levels, compared to healthy controls. Trophoblast or endothelial cells were exposed to low/high oxygen, and treated with MSC-conditioned media (MSC-CM) or a FKBPL overexpression plasmid. MAIN OUTCOME MEASURES: Preeclampsia risk stratification and diagnostic potential of FKBPL and CD44 were investigated. MSC treatment effects and FKBPL-CD44 signaling in trophoblast and endothelial cells were assessed. RESULTS: The CD44/FKBPL ratio was reduced in placenta and plasma following clinical diagnosis of preeclampsia. At 20 weeks of gestation, a high plasma CD44/FKBPL ratio was independently associated with the 2.3-fold increased risk of preeclampsia (odds ratio = 2.3, 95% confidence interval [CI] 1.03-5.23, P = 0.04). In combination with high mean arterial blood pressure (>82.5 mmHg), the risk further increased to 3.9-fold (95% CI 1.30-11.84, P = 0.016). Both hypoxia and MSC-based therapy inhibited FKBPL-CD44 signaling, enhancing cell angiogenesis. CONCLUSIONS: The FKBPL-CD44 pathway appears to have a central role in the pathogenesis of preeclampsia, showing promising utilities for early diagnostic and therapeutic purposes.

Hypercapnic acidosis induces mitochondrial dysfunction and impairs the ability of mesenchymal stem cells to promote distal lung epithelial repair.

Acute respiratory distress syndrome (ARDS) is a devastating disorder characterized by diffuse inflammation and edema formation. The main management strategy, low tidal volume ventilation, can be associated with the development of hypercapnic acidosis (HCA). Mesenchymal stem cells (MSCs) are a promising therapeutic candidate currently in early-phase clinical trials. The effects of HCA on the alveolar epithelium and capillary endothelium are not well established. The therapeutic efficacy of MSCs has never been reported in HCA. In the present study, we evaluated the effects of HCA on inflammatory response and reparative potential of the primary human small airway epithelial and lung microvasculature endothelial cells as well as on the capacity of bone marrow-derived MSCs to promote wound healing in vitro. We demonstrate that HCA attenuates the inflammatory response and reparative potential of primary human small airway epithelium and capillary endothelium and induces mitochondrial dysfunction. It was found that MSCs promote lung epithelial wound repair via the transfer of functional mitochondria; however, this proreparative effect of MSCs was lost in the setting of HCA. Therefore, HCA may adversely impact recovery from ARDS at the cellular level, whereas MSCs may not be therapeutically beneficial in patients with ARDS who develop HCA.-Fergie, N., Todd, N., McClements, L., McAuley, D., O'Kane, C., Krasnodembskaya, A. Hypercapnic acidosis induces mitochondrial dysfunction and impairs the ability of mesenchymal stem cells to promote distal lung epithelial repair.

Biotransformation of Silver Released from Nanoparticle Coated Titanium Implants Revealed in Regenerating Bone.

Antimicrobial silver nanoparticle coatings have attracted interest for reducing prosthetic joint infection. However, few studies report in vivo investigations of the biotransformation of silver nanoparticles within the regenerating tissue and its impact on bone formation. We present a longitudinal investigation of the osseointegration of silver nanoparticle-coated additive manufactured titanium implants in rat tibial defects. Correlative imaging at different time points using nanoscale secondary ion mass spectrometry, transmission electron microscopy (TEM), histomorphometry, and 3D X-ray microcomputed tomography provided quantitative insight from the nano- to macroscales. The quality and quantity of newly formed bone is comparable between the uncoated and silver coated implants. The newly formed bone demonstrates a trabecular morphology with bone being located at the implant surface, and at a distance, at two weeks. Nanoscale elemental mapping of the bone-implant interface showed that silver was present primarily in the osseous tissue and colocalized with sulfur. TEM revealed silver sulfide nanoparticles in the newly regenerated bone, presenting strong evidence that the previously in vitro observed biotransformation of silver to silver sulfide occurs in vivo.

Highly degradable porous melt-derived bioactive glass foam scaffolds for bone regeneration.

A challenge in using bioactive melt-derived glass in bone regeneration is to produce scaffolds with interconnected pores while maintaining the amorphous nature of the glass and its associated bioactivity. Here we introduce a method for creating porous melt-derived bioactive glass foam scaffolds with low silica content and report in vitro and preliminary in vivo data. The gel-cast foaming process was adapted, employing temperature controlled gelation of gelatin, rather than the in situ acrylic polymerisation used previously. To form a 3D construct from melt derived glasses, particles must be fused via thermal processing, termed sintering. The original Bioglass® 45S5 composition crystallises upon sintering, altering its bioactivity, due to the temperature difference between the glass transition temperature and the crystallisation onset being small. Here, we optimised and compared scaffolds from three glass compositions, ICIE16, PSrBG and 13-93, which were selected due to their widened sintering windows. Amorphous scaffolds with modal pore interconnect diameters between 100-150µm and porosities of 75% had compressive strengths of 3.4±0.3MPa, 8.4±0.8MPa and 15.3±1.8MPa, for ICIE16, PSrBG and 13-93 respectively. These porosities and compressive strength values are within the range of cancellous bone, and greater than previously reported foamed scaffolds. Dental pulp stem cells attached to the scaffold surfaces during in vitro culture and were viable. In vivo, the scaffolds were found to regenerate bone in a rabbit model according to X-ray micro tomography imaging. STATEMENT OF SIGNIFICANCE: This manuscript describes a new method for making scaffolds from bioactive glasses using highly bioactive glass compositions. The glass compositions have lower silica content that those that have been previously made into amorphous scaffolds and they have been designed to have similar network connectivity to that of the original (and commercially used) 45S5 Bioglass. The aim was to match Bioglass' bioactivity. The scaffolds retain the amorphous nature of bioactive glass while having an open pore structure and compressive strength similar to porous bone (the original 45S5 Bioglass crystallises during sintering, which can cause reduced bioactivity or instability). The new scaffolds showed unexpectedly rapid bone regeneration in a rabbit model.

Atomic Layer Deposition of a Silver Nanolayer on Advanced Titanium Orthopedic Implants Inhibits Bacterial Colonization and Supports Vascularized de Novo Bone Ingrowth.

Joint replacement surgery is associated with significant morbidity and mortality following infection with either methicillin-resistant Staphylococcus aureus (MRSA) or Staphylococcus epidermidis. These organisms have strong biofilm-forming capability in deep wounds and on prosthetic surfaces, with 103 -104 microbes resulting in clinically significant infections. To inhibit biofilm formation, we developed 3D titanium structures using selective laser melting and then coated them with a silver nanolayer using atomic layer deposition. On bare titanium scaffolds, S. epidermidis growth was slow but on silver-coated implants there were significant further reductions in both bacterial recovery (p < 0.0001) and biofilm formation (p < 0.001). MRSA growth was similarly slow on bare titanium scaffolds and not further affected by silver coating. Ultrastructural examination and viability assays using either human bone or endothelial cells, demonstrated strong adherence and growth on titanium-only or silver-coated implants. Histological, X-ray computed microtomographic, and ultrastructural analyses revealed that silver-coated titanium scaffolds implanted into 2.5 mm defects in rat tibia promoted robust vascularization and conspicuous bone ingrowth. We conclude that nanolayer silver of titanium implants significantly reduces pathogenic biofilm formation in vitro, facilitates vascularization and osseointegration in vivo making this a promising technique for clinical orthopedic applications.

A correlative imaging based methodology for accurate quantitative assessment of bone formation in additive manufactured implants.

A correlative imaging methodology was developed to accurately quantify bone formation in the complex lattice structure of additive manufactured implants. Micro computed tomography (µCT) and histomorphometry were combined, integrating the best features from both, while demonstrating the limitations of each imaging modality. This semi-automatic methodology registered each modality using a coarse graining technique to speed the registration of 2D histology sections to high resolution 3D µCT datasets. Once registered, histomorphometric qualitative and quantitative bone descriptors were directly correlated to 3D quantitative bone descriptors, such as bone ingrowth and bone contact. The correlative imaging allowed the significant volumetric shrinkage of histology sections to be quantified for the first time (~15 %). This technique demonstrated the importance of location of the histological section, demonstrating that up to a 30 % offset can be introduced. The results were used to quantitatively demonstrate the effectiveness of 3D printed titanium lattice implants.