Anti-cancer management of head and neck cancers and oral microbiome-what can we clinically obtain?

Head and neck squamous cell carcinoma (HNSCC) exhibits significant genetic heterogeneity and primarily concerns the oral cavity and oropharynx. These cancers occur more frequently in men with a 5-year survival rate below 50%. Major risk factors include human papilloma virus (HPV) (notably type 16), Epstein-Barr virus, tobacco, alcohol, and poor oral hygiene with approximately 4.5% of global cancers linked to HPV. Notably, differences in the microbiome between healthy individuals and patients with head and neck cancers (HNCs) have been identified. Recent studies highlight the significance of certain oral microbes in risk assessment and the potential of the microbiome as a biomarker for HNCs. Additionally, role of the microbiome in metastasis has been acknowledged. Treatment for HNCs includes local methods, such as surgery and radiotherapy, and systemic approaches, such as immunotherapy. Numerous side effects accompany these treatments. Emerging research suggests the beneficial role of preoperative immunonutrition and probiotics in patient outcomes, emphasizing the influence of the microbiome on treatment efficacy. This review explores the reciprocal effects of HNC treatment and the gut microbiome using radiotherapy, brachytherapy, surgery, immunotherapy, and chemotherapy.

Coordination Chemistry of Sulfur-Containing Bifunctional Chelators: Toward in Vivo Stabilization of 64Cu PET Imaging Agents for Alzheimer's Disease.

The broader utilization of 64Cu positron emission tomography (PET) imaging agents has been hindered by the unproductive demetalation induced by bioreductants. To advance the development of 64Cu-based PET imaging tracers for Alzheimer's Disease (AD), there is a need for novel ligand design strategies. In this study, we developed sulfur-containing dithiapyridinophane (N2S2) bifunctional chelators (BFCs) as well as all nitrogen-based diazapyridinophane (N4) BFCs to compare their abilities to chelate Cu and target Aß aggregates. Through spectrophotometric titrations and electrochemical measurements, we have demonstrated that the N2S2-based BFCs exhibit >10 orders of magnitude higher binding affinity toward Cu(I) compared to their N4-based counterparts, while both types of BFCs exhibit high stability constants toward Cu(II). Notably, solid state structures for both Cu(II) and Cu(I) complexes supported by the two ligand frameworks were obtained, providing molecular insights into their copper chelating abilities. Aß binding experiments were conducted to study the structure-affinity relationship, and fluorescence microscopy imaging studies confirmed the selective labeling of the BFCs and their copper complexes. Furthermore, we investigated the potential of these ligands for the 64Cu-based PET imaging of AD through radiolabeling and autoradiography studies. We believe our findings provide molecular insights into the design of bifunctional Cu chelators that can effectively stabilize both Cu(II) and Cu(I) and, thus, can have significant implications for the development of 64Cu PET imaging as a diagnostic tool for AD.

In Vitro and In Vivo Imaging-Based Evaluation of Doxorubicin Anticancer Treatment in Combination with the Herbal Medicine Black Cohosh.

As a substitution for hormone replacement therapy, many breast cancer patients use black cohosh (BC) extracts in combination with doxorubicin (DOX)-based chemotherapy. In this study, we evaluated the viability and survival of BC- and DOX-treated MCF-7 cells. A preclinical model of MCF-7 xenografts was used to determine the influence of BC and DOX administration on tumor growth and metabolism. The number of apoptotic cells after incubation with both DOX and BC was significantly increased (~100%) compared to the control. Treatment with DOX altered the potential of MCF-7 cells to form colonies; however, coincubation with BC did not affect this process. In vivo, PET-CT imaging showed that combined treatment of DOX and BC induced a significant reduction in both metabolic activity (29%) and angiogenesis (32%). Both DOX and BC treatments inhibited tumor growth by 20% and 12%, respectively, and combined by 57%, vs. control. We successfully demonstrated that BC increases cytotoxic effects of DOX, resulting in a significant reduction in tumor size. Further studies regarding drug transport and tumor growth biomarkers are necessary to establish the underlying mechanism and potential clinical use of BC in breast cancer patients.

Receptor for advanced glycation end-products: Biological significance and imaging applications.

The receptor for advanced glycation end-products (RAGE or AGER) is a transmembrane, immunoglobulin-like receptor that, due to its multiple isoform structures, binds to a diverse range of endo- and exogenous ligands. RAGE activation caused by the ligand binding initiates a cascade of complex pathways associated with producing free radicals, such as reactive nitric oxide and oxygen species, cell proliferation, and immunoinflammatory processes. The involvement of RAGE in the pathogenesis of disorders such as diabetes, inflammation, tumor progression, and endothelial dysfunction is dictated by the accumulation of advanced glycation end-products (AGEs) at pathologic states leading to sustained RAGE upregulation. The involvement of RAGE and its ligands in numerous pathologies and diseases makes RAGE an interesting target for therapy focused on the modulation of both RAGE expression or activation and the production or exogenous administration of AGEs. Despite the known role that the RAGE/AGE axis plays in multiple disease states, there remains an urgent need to develop noninvasive, molecular imaging approaches that can accurately quantify RAGE levels in vivo that will aid in the validation of RAGE and its ligands as biomarkers and therapeutic targets. This article is categorized under: Diagnostic Tools > In Vivo Nanodiagnostics and Imaging Diagnostic Tools > Biosensing.

RAGE as a Novel Biomarker for Prostate Cancer: A Systematic Review and Meta-Analysis.

The receptor for advanced glycation end-products (RAGE) has been implicated in driving prostate cancer (PCa) growth, aggression, and metastasis through the fueling of chronic inflammation in the tumor microenvironment. This systematic review and meta-analysis summarizes and analyzes the current clinical and preclinical data to provide insight into the relationships among RAGE levels and PCa, cancer grade, and molecular effects. A multi-database search was used to identify original clinical and preclinical research articles examining RAGE expression in PCa. After screening and review, nine clinical and six preclinical articles were included. The associations of RAGE differentiating benign prostate hyperplasia (BPH) or normal prostate from PCa and between tumor grades were estimated using odds ratios (ORs) and associated 95% confidence intervals (CI). Pooled estimates were calculated using random-effect models due to study heterogeneity. The clinical meta-analysis found that RAGE expression was highly likely to be increased in PCa when compared to BPH or normal prostate (OR: 11.3; 95% CI: 4.4-29.1) and that RAGE was overexpressed in high-grade PCa when compared to low-grade PCa (OR: 2.5; 95% CI: 1.8-3.4). In addition, meta-analysis estimates of preclinical studies performed by albatross plot generation found robustly positive associations among RAGE expression/activation and PCa growth and metastatic potential. This review demonstrates that RAGE expression is strongly tied to PCa progression and can serve as an effective diagnostic target to differentiate between healthy prostate, low-grade PCa, and high-grade PCa, with potential theragnostic applications.

Proteomic and Metabolomic Changes in Psoriasis Preclinical and Clinical Aspects.

Skin diseases such as psoriasis (Ps) and psoriatic arthritis (PsA) are immune-mediated inflammatory diseases. Overlap of autoinflammatory and autoimmune conditions hinders diagnoses and identifying personalized patient treatments due to different psoriasis subtypes and the lack of verified biomarkers. Recently, proteomics and metabolomics have been intensively investigated in a broad range of skin diseases with the main purpose of identifying proteins and small molecules involved in the pathogenesis and development of the disease. This review discusses proteomics and metabolomics strategies and their utility in research and clinical practice in psoriasis and psoriasis arthritis. We summarize the studies, from in vivo models conducted on animals through academic research to clinical trials, and highlight their contribution to the discovery of biomarkers and targets for biological drugs.

Molecular Imaging and Nanotechnology-Emerging Tools in Diagnostics and Therapy.

Personalized medicine is emerging as a new goal in the diagnosis and treatment of diseases. This approach aims to establish differences between patients suffering from the same disease, which allows to choose the most effective treatment. Molecular imaging (MI) enables advanced insight into molecule interactions and disease pathology, improving the process of diagnosis and therapy and, for that reason, plays a crucial role in personalized medicine. Nanoparticles are widely used in MI techniques due to their size, high surface area to volume ratio, and multifunctional properties. After conjugation to specific ligands and drugs, nanoparticles can transport therapeutic compounds directly to their area of action and therefore may be used in theranostics-the simultaneous implementation of treatment and diagnostics. This review summarizes different MI techniques, including optical imaging, ultrasound imaging, magnetic resonance imaging, nuclear imaging, and computed tomography imaging with theranostics nanoparticles. Furthermore, it explores the potential use of constructs that enables multimodal imaging and track diseases in real time.

Endothelial Dysfunction Driven by Hypoxia-The Influence of Oxygen Deficiency on NO Bioavailability.

Cardiovascular diseases (CVDs) are the leading cause of death worldwide. The initial stage of CVDs is characterized by endothelial dysfunction, defined as the limited bioavailability of nitric oxide (NO). Thus, any factors that interfere with the synthesis or metabolism of NO in endothelial cells are involved in CVD pathogenesis. It is well established that hypoxia is both the triggering factor as well as the accompanying factor in cardiovascular disease, and diminished tissue oxygen levels have been reported to influence endothelial NO bioavailability. In endothelial cells, NO is produced by endothelial nitric oxide synthase (eNOS) from L-Arg, with tetrahydrobiopterin (BH4) as an essential cofactor. Here, we discuss the mechanisms by which hypoxia affects NO bioavailability, including regulation of eNOS expression and activity. What is particularly important is the fact that hypoxia contributes to the depletion of cofactor BH4 and deficiency of substrate L-Arg, and thus elicits eNOS uncoupling-a state in which the enzyme produces superoxide instead of NO. eNOS uncoupling and the resulting oxidative stress is the major driver of endothelial dysfunction and atherogenesis. Moreover, hypoxia induces impairment in mitochondrial respiration and endothelial cell activation; thus, oxidative stress and inflammation, along with the hypoxic response, contribute to the development of endothelial dysfunction.

Molecularly targeted nanoparticles: an emerging tool for evaluation of expression of the receptor for advanced glycation end products in a murine model of peripheral artery disease.

BACKGROUND: Molecular imaging with molecularly targeted probes is a powerful tool for studying the spatio-temporal interactions between complex biological processes. The pivotal role of the receptor for advanced glycation end products (RAGE), and its involvement in numerous pathological processes, aroused the demand for RAGE-targeted imaging in various diseases. In the present study, we evaluated the use of a diagnostic imaging agent for RAGE quantification in an animal model of peripheral artery disease, a multimodal dual-labeled probe targeted at RAGE (MMIA-CML). METHODS: PAMAM dendrimer was conjugated with Nε-carboxymethyl-lysine (CML) modified albumin to synthesize the RAGE-targeted probe. A control untargeted agent carried native non-modified human albumin (HSA). Bifunctional p-SCN-Bn-NOTA was used to conjugate the 64Cu radioisotope. Surgical right femoral artery ligation was performed on C57BL/6 male mice. One week after femoral artery ligation, mice were injected with MMIA-CML or MMIA-HSA labeled with 64Cu radioisotope and 60 min later in vivo microPET-CT imaging was performed. Immediately after PET imaging studies, the murine hindlimb muscle tissues were excised and prepared for gene and protein expression analysis. RAGE gene and protein expression was assessed using real-time qPCR and Western blot technique respectively. To visualize RAGE expression in excised tissues, microscopic fluorescence imaging was performed using RAGE-specific antibodies and RAGE-targeted and -control MMIA. RESULTS: Animals subjected to PET imaging exhibited greater MMIA-CML uptake in ischemic hindlimbs than non-ischemic hindlimbs. We observed a high correlation between fluorescent signal detection and radioactivity measurement. Significant RAGE gene and protein overexpression were observed in ischemic hindlimbs compared to non-ischemic hindlimbs at one week after surgical ligation. Fluorescence microscopic staining revealed significantly increased uptake of RAGE-targeted nanoparticles in both ischemic and non-ischemic muscle tissues compared to the control probe but at a higher level in ischemic hindlimbs. Ischemic tissue exhibited explicit RAGE dyeing following anti-RAGE antibody and high colocalization with the MMIA-CML targeted at RAGE. CONCLUSIONS: The present results indicate increased expression of RAGE in the ischemic hindlimb and enable the use of multimodal nanoparticles in both in vitro and in vivo experimental models, creating the possibility for imaging structural and functional changes with a RAGE-targeted tracer.

Pathogenesis of psoriasis in the "omic" era. Part IV. Epidemiology, genetics, immunopathogenesis, clinical manifestation and treatment of psoriatic arthritis.

Psoriatic arthritis (PsA) is a chronic, progressive, inflammatory arthropathy associated with psoriasis as well as a complex pathogenesis. Genetic and environmental factors trigger the development of the immune-mediated auto-inflammatory response in different sites: skin, bone marrow, entheses and synovial tissues. Studies of the last two decades have changed the view of PsA from a mild, non-progressive arthritis to an inflammatory systemic disease with serious health consequences, not only associated with joint dysfunction, but also with an increased risk of cardiovascular disease and socioeconomic consequences with significantly reduced quality of life. The joint damage starts early in the course of the disease, thus early recognition and treatment with modern biological treatments, which may modify the natural history and slow down progression of this debilitating disease, is essential for the patient long-term outcome.

Pathogenesis of psoriasis in the "omic" era. Part III. Metabolic disorders, metabolomics, nutrigenomics in psoriasis.

Psoriasis is a systemic disease that is strictly connected with metabolic disorders (insulin resistance, atherogenic dyslipidemia, arterial hypertension, and cardiovascular diseases). It occurs more often in patients with a more severe course of the disease. Obesity is specially an independent risk factor and it is associated with a worse treatment outcome because of the high inflammatory activity of visceral fatty tissue and the production of inflammatory mediators involved in the development of both psoriasis and metabolic disorders. However, in psoriasis the activation of the Th17/IL-17 and the abnormalities in the Th17/Treg balance axis are observed, but this pathomechanism does not fully explain the frequent occurrence of metabolic disorders. Therefore, there is a need to look for better biomarkers in the diagnosis, prognosis and monitoring of concomitant disorders and therapeutic effects in psoriasis. In addition, the education on the use of a proper diet as a prophylaxis for the development of the above disorders is an important element of holistic care for a patient with psoriasis. Diet may affect gene expression due to epigenetic modification which encompasses interactions of environment, nutrition and diseases. Patients with psoriasis should be advised to adopt proper diet and dietician support.

Genetic and Epigenetic Aspects of Atopic Dermatitis.

Atopic dermatitis is a heterogeneous disease, in which the pathogenesis is associated with mutations in genes encoding epidermal structural proteins, barrier enzymes, and their inhibitors; the role of genes regulating innate and adaptive immune responses and environmental factors inducing the disease is also noted. Recent studies point to the key role of epigenetic changes in the development of the disease. Epigenetic modifications are mainly mediated by DNA methylation, histone acetylation, and the action of specific non-coding RNAs. It has been documented that the profile of epigenetic changes in patients with atopic dermatitis (AD) differs from that observed in healthy people. This applies to the genes affecting the regulation of immune response and inflammatory processes, e.g., both affecting Th1 bias and promoting Th2 responses and the genes of innate immunity, as well as those encoding the structural proteins of the epidermis. Understanding of the epigenetic alterations is therefore pivotal to both create new molecular classifications of atopic dermatitis and to enable the development of personalized treatment strategies.

Pathogenesis of psoriasis in the "omic" era. Part II. Genetic, genomic and epigenetic changes in psoriasis.

Psoriasis is a multifactorial disease in which genetic, environmental and epigenetic factors regulating gene expression play a key role. In the "genomic era", genome-wide association studies together with target genotyping platforms performed in different ethnic populations have found more than 50 genetic susceptible markers associated with the risk of psoriasis which have been identified so far. Up till now, the strongest association with the risk of the disease has been proved for HLA-C*06 gene. The majority of other psoriasis risk SNPs are situated near the genes encoding molecules involved in adaptive and innate immunity, and skin barrier function. Many contemporary studies indicate that the epigenetic changes: histone modification, promoter methylations, long non-coding and micro-RNA hyperexpression are considered as factors contributing to psoriasis pathogenesis as they regulate abnormal keratinocyte differentiation and proliferation, aberrant keratinocytes - inflammatory cells communication, neoangiogenesis and chronic inflammation. The circulating miRNAs detected in the blood may become specific markers in the diagnosis, prognosis and response to the treatment of the disease. The inhibition of expression in selected miRNAs may be a new promising therapy option for patients with psoriasis.

Pathogenesis of psoriasis in the "omic" era. Part I. Epidemiology, clinical manifestation, immunological and neuroendocrine disturbances.

Psoriasis is a common, chronic, inflammatory, immune-mediated skin disease affecting about 2% of the world's population. According to current knowledge, psoriasis is a complex disease that involves various genes and environmental factors, such as stress, injuries, infections and certain medications. The chronic inflammation of psoriasis lesions develops upon epidermal infiltration, activation, and expansion of type 1 and type 17 Th cells. Despite the enormous progress in understanding the mechanisms that cause psoriasis, the target cells and antigens that drive pathogenic T cell responses in psoriatic lesions are still unproven and the autoimmune basis of psoriasis still remains hypothetical. However, since the identification of the Th17 cell subset, the IL-23/Th17 immune axis has been considered a key driver of psoriatic inflammation, which has led to the development of biologic agents that target crucial elements of this pathway. Here we present the current understanding of various aspects in psoriasis pathogenesis.

Quantitative imaging of the receptor for advanced glycation end-products in prostate cancer.

PURPOSE: Current screening and monitoring of prostate cancer (PCa) is insufficient, producing inaccurate diagnoses. Presence of the receptor for advanced glycation end-products (RAGE) is associated with signature characteristics of PCa development such as cell proliferation, anchorage-independent growth, angiogenesis, migration, invasion, and poor patient survival. Therefore, we developed a preclinical multimodal imaging strategy targeted at RAGE to diagnose and monitor PCa. METHODS: In this work, RAGE-targeted multimodal nanoparticles (64Cu-Cy5-G4-CML) were synthesized and rendered functional for nuclear and optical imaging using previously established methods. The probe's binding affinity and targeting specificity was assessed in androgen-dependent (LNCaP) and androgen-independent (DU145) prostate cancer cells using flow cytometry and confocal microscopy. In vivo PET-CT imaging was used to evaluate RAGE levels in DU145 and LNCaP xenograft models in mice. Then, tumors were excised post-imaging for histological staining and autoradiography to further assess RAGE levels and targeting efficiency of the tracer. Finally, RAGE levels from human PCa samples of varying Gleason Scores were evaluated using Western blot and immunohistochemical staining. RESULTS: PCa cell culture studies confirmed adequate RAGE-targeting with 64Cu-Cy5-G4-CML with KD between 360 and 540 nM as measured by flow cytometry. In vivo PET-CT images of PCa xenografts revealed favorable kinetics, rapid blood clearance, and a non-homogenous, enhanced uptake in tumors, which varied based on cell type and tumor size with mean uptake between 0.5 and 1.4%ID/g. RAGE quantification of human samples confirmed increased RAGE uptake corresponding to increased Gleason scoring. CONCLUSIONS: Our study has shown that RAGE-targeted cancer imaging is feasible and could significantly impact PCa management.

Imaging the Landmarks of Vascular Recovery.

Background: Peripheral arterial disease (PAD) is a major worldwide health concern. Since the late 1990s therapeutic angiogenesis has been investigated as an alternative to traditional PAD treatments. Although positive preclinical results abound in the literature, the outcomes of human clinical trials have been discouraging. Among the challenges the field has faced has been a lack of standardization of the timings and measures used to validate new treatment approaches. Methods: In order to study the spatiotemporal dynamics of both perfusion and neovascularization in mice subjected to surgically-induced hindlimb ischemia (n= 30), we employed three label-free imaging modalities (a novel high-sensitivity ultrasonic Power Doppler methodology, laser speckle contrast, and photoacoustic imaging), as well as a tandem of radio-labeled molecular probes, 99mTc-NC100692 and 99mTc-BRU-5921 respectively, designed to detect two key modulators of angiogenic activity, αVß3 and HIF-1α , via scintigraphic imaging. Results: The multimodal imaging strategy reveals a set of "landmarks"-key physiological and molecular events in the healing process-that can serve as a standardized framework for describing the impact of emerging PAD treatments. These landmarks span the entire process of neovascularization, beginning with the rapid decreases in perfusion and oxygenation associated with ligation surgery, extending through pro-angiogenic changes in gene expression driven by the master regulator HIF-1α , and ultimately leading to complete functional revascularization of the affected tissues. Conclusions: This study represents an important step in the development of multimodal non-invasive imaging strategies for vascular research; the combined results offer more insight than can be gleaned through any of the individual imaging methods alone. Researchers adopting similar imaging strategies and will be better able to describe changes in the onset, duration, and strength of each of the landmarks of vascular recovery, yielding greater biological insight, and enabling more comprehensive cross-study comparisons. Perhaps most important, this study paves the road for more efficient translation of PAD research; emerging experimental treatments can be more effectively assessed and refined at the preclinical stage, ultimately leading to better next-generation therapies.

Streptococcus pneumoniae Elaborates Persistent and Prolonged Competent State during Pneumonia-Derived Sepsis.

The competence regulon of pneumococcus regulates both genetic transformation and virulence. However, competence induction during host infection has not been examined. By using the serotype 2 strain D39, we transcriptionally fused the firefly luciferase (luc) to competence-specific genes and spatiotemporally monitored the competence development in a mouse model of pneumonia-derived sepsis. In contrast to the universally reported short transient burst of competent state in vitro, the naturally developed competent state was prolonged and persistent during pneumonia-derived sepsis. The competent state began at approximately 20 h postinfection (hpi) and facilitated systemic invasion and sepsis development and progressed in different manners. In some mice, acute pneumonia quickly led to sepsis and death, accompanied by increasing intensity of the competence signal. In the remaining mice, pneumonia lasted longer, with the competence signal decreasing at first but increasing as the infection became septic. The concentration of pneumococcal inoculum (1 × 106 to 1 × 108 CFU/mouse) and postinfection lung bacterial burden did not appreciably impact the kinetics of competence induction. Exogenously provided competence stimulating peptide 1 (CSP1) failed to modulate the onset kinetics of competence development in vivo The competence shutoff regulator DprA was highly expressed during pneumonia-derived sepsis but failed to turn off the competent state in mice. Competent D39 bacteria propagated the competence signal through cell-to-cell contact rather than the classically described quorum-sensing mechanism. Finally, clinical pneumococcal strains of different serotypes were also able to develop natural competence during pneumonia-derived sepsis.

Multimodal imaging of the receptor for advanced glycation end-products with molecularly targeted nanoparticles.

The receptor for advanced glycation end-products (RAGE) is central to multiple disease states, including diabetes-related conditions such as peripheral arterial disease (PAD). Despite RAGE's importance in these pathologies, there remains a need for a molecular imaging agent that can accurately assess RAGE levels in vivo. Therefore, we have developed a multimodal nanoparticle-based imaging agent targeted at RAGE with the well-characterized RAGE ligand, carboxymethyllysine (CML)-modified human serum albumin (HSA). Methods: A multimodal tracer (64Cu-Rho-G4-CML) was developed using a generation-4 (G4) polyamidoamine (PAMAM) dendrimer, conjugated with both rhodamine and copper-64 (64Cu) chelator (NOTA) for optical and PET imaging, respectively. First, 64Cu-Rho-G4-CML and its non-targeted analogue (64Cu-Rho-G4-HSA) were evaluated chemically using techniques such as dynamic light scattering (DLS), electron microscopy and nuclear magnetic resonance (NMR). The tracers' binding capabilities were examined at the cellular level and optimized using live and fixed HUVEC cells grown in 5.5-30 mM glucose, followed by in vivo PET-CT imaging, where the probes' kinetics, biodistribution, and RAGE targeting properties were examined in a murine model of hindlimb ischemia. Finally, histological assessment of RAGE levels in both ischemic and non-ischemic tissues was performed. Conclusions: Our RAGE-targeted probe demonstrated an average size of 450 nm, a Kd of 340-390 nM, rapid blood clearance, and a 3.4 times greater PET uptake in ischemic RAGE-expressing hindlimbs than their non-ischemic counterpart. We successfully demonstrated increased RAGE expression in a murine model of hindlimb ischemia and the feasibility for non-invasive examination of cellular, tissue, and whole-body RAGE levels with a molecularly targeted tracer.

Evaluation of a dimeric-cRGD peptide for targeted PET-CT imaging of peripheral angiogenesis in diabetic mice.

The α V ß3 integrin plays an important role in many physiological functions and pathological disorders. α V ß3 is minimally expressed in normal quiescent endothelial cells, but significantly upregulated during neovascularization. In this study, we evaluated a 64Cu-labeled dimeric cRGD tracer targeted at α V ß3 integrin and report its applicability to assess peripheral angiogenesis in diabetes mellitus (DM). We established a murine model of type-1 DM characterized by elevated glucose, glycated serum protein (GSP), and glycated hemoglobin A1c (HbA1c). We demonstrated that our imaging probe is specific to α V ß3 integrin under both normo- and hyperglycemic conditions. We found that the analysis of in vivo PET-CT images correlated well with gamma well counting (GWC). Both GWC and PET-CT imaging demonstrated increased uptake of 64Cu-NOTA-PEG4-cRGD2 in the ischemic hindlimb in contrast to non-ischemic control. GWC of the distal ischemic tissue from DM mice showed significantly lower probe accumulation than in non-DM mice. The immunofluorescence staining of the ischemic tissues showed a 3-fold reduction in CD31 and 4-fold reduction in the α V ß3 expression in DM vs. non-DM animals. In conclusion, we successfully demonstrated that diabetes-associated reductions in peripheral angiogenesis can be non-invasively detected with PET-CT imaging using targeted dimeric-cRGD probe.

The impact of mechanically stimulated muscle-derived stromal cells on aged skeletal muscle.

Perivascular stromal cells, including mesenchymal stem/stromal cells (MSCs), secrete paracrine factor in response to exercise training that can facilitate improvements in muscle remodeling. This study was designed to test the capacity for muscle-resident MSCs (mMSCs) isolated from young mice to release regenerative proteins in response to mechanical strain in vitro, and subsequently determine the extent to which strain-stimulated mMSCs can enhance skeletal muscle and cognitive performance in a mouse model of uncomplicated aging. Protein arrays confirmed a robust increase in protein release at 24h following an acute bout of mechanical strain in vitro (10%, 1Hz, 5h) compared to non-strain controls. Aged (24month old), C57BL/6 mice were provided bilateral intramuscular injection of saline, non-strain control mMSCs, or mMSCs subjected to a single bout of mechanical strain in vitro (4×104). No significant changes were observed in muscle weight, myofiber size, maximal force, or satellite cell quantity at 1 or 4wks between groups. Peripheral perfusion was significantly increased in muscle at 4wks post-mMSC injection (p<0.05), yet no difference was noted between control and preconditioned mMSCs. Intramuscular injection of preconditioned mMSCs increased the number of new neurons and astrocytes in the dentate gyrus of the hippocampus compared to both control groups (p<0.05), with a trend toward an increase in water maze performance noted (p=0.07). Results from this study demonstrate that acute injection of exogenously stimulated muscle-resident stromal cells do not robustly impact aged muscle structure and function, yet increase the survival of new neurons in the hippocampus.