Advanced Regenerative Techniques Based on Dental Pulp Stem Cells for the Treatment of Periodontal Disease

Recent progress in periodontology intended to reduce the risk represented by periodontal disease for systemic disorders and general human health condition. In this chapter, we overview the advantages and limitations of current techniques based on occlusive membranes for periodontal regeneration. Special emphasis is paid to advanced techniques using stem cells from dental pulp for the regeneration of bone defects caused by the chronic periodontal disease. Stem cells isolation, in vitro expansion and characterization techniques are presented. Therapeutic strategies of stem cells delivery using natural polymeric carriers are discussed. Stem cell-scaffold constructs application in bone tissue engineering is proposed, taking into account the marked decline of healing, and regenerative processes in elderly individuals. Future researchers envisage multiple effects of engineered constructs with antimicrobial, anti-inflammatory, and regenerative activity for periodontal treatment

Uklanjanje slobodnih radikala, postizanje redoks ravnoteže i cijeljenje rana pomoću bioaktivnih peptida dobivenih hidrolizom kolagena iz kože bijelog glavaša (Hypophthalmichthys molitrix) potpomognutom proteinazom K

Research background. Various protocols for enzymatic hydrolysis of fish by-products are increasingly tested to ensure value-added products with functional and biological properties important for food, cosmetic and medical applications. In addition, they attempt to minimize waste from industrial processing and environmental requirements. This study aims to establish an efficient protocol based on two-step enzymatic hydrolysis of freshwater fish skin and to evaluate the effect of resulting bioactive peptides on free radical scavenging, redox balance and regulation of fibroblast proliferation and migration. Experimental approach. Pepsin-soluble collagen extracted from silver carp (Hypophthalmichthys molitrix) skin was hydrolyzed by proteinase K at specific sites under controlled conditions. The molecular mass of ultrafiltration permeate was determined by gradient electrophoresis and gel filtration chromatography. The biological activity of intermediate and small size bioactive peptides was evaluated in experimental models in vitro mimicking oxidative stress and skin wound conditions. Results and conclusions. Extracted fish collagen was hydrolysed using proteinase K, the most efficient enzyme for the cleavage of the primary structure of the molecule, as previously found in silico. Established optimal conditions increased the enzyme specificity and the process yield. Bioactive peptides exerted significantly higher scavenging activity on free stable radicals and hydroxyl radicals often found in vivo, compared to fish collagen. They stimulated fibroblast metabolism in a dose-dependent manner and up-regulated cell migration in a scratch wound model. Pretreatment of fibroblasts with induced oxidative stress using optimal concentrations of fish peptides prevented the increase of reactive oxygen species production. In conclusion, bioactive peptides from carp skin demonstrated valuable properties of maintaining redox balance and skin wound healing process improvement, which indicated further potential applications in the development of pharmaceutical and nutraceutical formulations. Novelty and scientific contribution. In this study the enzymatic hydrolysis was applied to isolated protein, in contrast to previous studies using waste tissue with variable composition. Recovered bioactive peptides acted not only as antioxidant agents, but also as regulators of oxidative stress and wound healing processes in skin cell models. Their nutritional and cosmetic application is recommended in novel formulations fighting skin ageing phenomena.Pozadina istraživanja. U posljednje se vrijeme sve više ispituju postupci enzimske hidrolize nusproizvoda obrade ribe radi dobivanja proizvoda obogaćenih funkcionalnim i biološkim svojstvima važnim u proizvodnji hrane, kozmetike i medicinskih preparata. Osim toga, na taj se način smanjuju količina otpada industrijske proizvodnje i pritisak na okoliš. Svrha je ovoga rada bila osmisliti učinkoviti postupak dvostupanjske enzimske hidrolize kože slatkovodne ribe i ispitati učinak dobivenih bioaktivnih peptida na uklanjanje slobodnih radikala, redoks ravnotežu, te proliferaciju i migraciju fibroblasta. Eksperimentalni pristup. Kolagen razgradljiv pomoću pepsina izoliran je iz kože bijelog glavaša (Hypoph¬thalmichthys molitrix) i hidroliziran u kontroliranim uvjetima pomoću proteinaze K. Molekulska masa permeata dobivenog ultrafiltracijom određena je pomoću elektroforeze s gradijentom i gel-filtracijske kromatografije. Biološka aktivnost peptida srednje i male veličine ispitana je in vitro simulacijom oksidacijskog stresa i oštećenja kože. Rezultati i zaključci. Riblji kolagen je hidroliziran pomoću proteinaze K, najučinkovitijeg enzima koji cijepa primarnu strukturu molekule, što je potvrđeno prethodnim ispitivanjem in silico. Pri optimalnim uvjetima povećali su se specifičnost enzima i prinos reakcije. U usporedbi s riblijm kolagenom, bioaktivni su peptidi imali bitno veću sposobnost uklanjanja slobodnih i hidroksilnih radikala, često prisutnih in vivo. Ovisno o koncentraciji, stimulirali su metabolizam fibroblasta i migraciju stanica pri cijeljenju modelne rane. Prethodna obrada fibroblasta optimalnim koncentracijama ribljih peptida spriječila je porast sinteze reaktivnih spojeva kisika pri induciranju oksidacijskog stresa. Možemo zaključiti da bioaktivni peptidi kože bijelog glavaša imaju važna svojstva, kao što su održavanje redoks ravnoteže i pospješivanje cijeljenja rana, što upućuje na njihovu moguću primjenu u razvoju farmaceutika i nutraceutika. Novina i znanstveni doprinos. U ovom je radu provedena enzimska hidroliza izoliranog proteina, za razliku od dosadašnjih istraživanja u kojima je korišten otpad različitog sastava. Dobiveni bioaktivni peptidi nisu djelovali samo kao antioksidacijski agensi, već i kao regulatori oksidacijskog stresa i cijeljenja kože u staničnom modelu. Zbog svojih svojstava mogu se preporučiti za primjenu u proizvodnji kozmetičkih pripravaka koji suzbijaju starenje kože

Preparation and Characterization of a Collagen-Liposome-Chondroitin Sulfate Matrix with Potential Application for Inflammatory Disorders Treatment

Smart drug delivery systems with controllable properties play an important role in targeted therapy and tissue regeneration. The aim of our study was the preparation and in vitro evaluation of a collagen (Col) matrix embedding a liposomal formulation of chondroitin sulfate (L-CS) for the treatment of inflammatory disorders. Structural studies using Oil Red O specific staining for lipids and scanning electron microscopy showed an alveolar network of nanosized Col fibrils decorated with deposits of L-CS at both periphery and inner of the matrix. The porosity and density of Col-L-CS matrix were similar to those of Col matrix, while its mean pore size and biodegradability had significantly higher and lower values (P<0.05), respectively. In vitro cytotoxicity assays showed that the matrix system induced high cell viability and stimulated cell metabolism in L929 fibroblast cell culture. Light and electron micrographs of the cell-matrix construct showed that cells clustered into the porous structure at 72 h of cultivation. In vitro diffusion test indicated that the quantity of released CS was significantly lower (P<0.05) after embedment of L-CS within Col matrix. All these results indicated that the biocompatible and biodegradable Col-L-CS matrix might be a promising delivery system for local treatment of inflamed site

A pilot clinical trial of intravesical mitomycin-C and external deep pelvic hyperthermia for non-muscle-invasive bladder cancer.

PURPOSE: This paper aims to evaluate the safety and heating efficiency of external deep pelvic hyperthermia combined with intravesical mitomycin C (MMC) as a novel therapy for non-muscle-invasive bladder cancer (NMIBC). MATERIALS AND METHODS: We enrolled subjects with bacillus Calmette-Guérin (BCG) refractory NMIBC to an early phase clinical trial of external deep pelvic hyperthermia (using a BSD-2000 device) combined with MMC. Bladders were heated to 42 °C for 1 h during intravesical MMC treatment. Treatments were given weekly for 6 weeks, then monthly for 4 months. Heating parameters, treatment toxicity, and clinical outcomes were systematically measured. RESULTS: Fifteen patients were enrolled on the clinical trial. Median age was 66 years and 87% were male. Median European Organisation for Research and Treatment of Cancer (EORTC) recurrence and progression scores were 6 and 8, respectively. The full treatment course was attained in 73% of subjects. Effective bladder heating was possible in all but one patient who could not tolerate the supine position due to lung disease. Adverse events were all minor (grade 2 or less) and no systemic toxicity was observed. The most common adverse effects were Foley catheter pain (40%), abdominal discomfort (33%), chemical cystitis symptoms (27%), and abdominal skin swelling (27%). With a median follow-up of 3.18 years, 67% experienced another bladder cancer recurrence (none were muscle invasive) and 13% experienced an upper tract recurrence. CONCLUSIONS: External deep pelvic hyperthermia using the BSD-2000 device is a safe and reproducible method of heating the bladder in patients undergoing intravesical MMC. The efficacy of this treatment modality should be explored further in clinical trials

Establishing Relevant ADC-based Texture Analysis Metrics for Quantifying Early Treatment-Induced Changes in Head and Neck Squamous Cell Carcinomas

Purpose: The purpose of this study is to identify which texture analysis metrics calculated from apparent diffusion coefficient (ADC) maps from patients with head and neck squamous cell carcinomas (HNSCC) provide quantifiable measures of tumor physiology changes. We discerned which imaging metrics were relevant using baseline agreement and variations during early treatment. Methods: For selective patients with stages II-IV HNSCC, ADC maps were generated from two baselines, taken 1 week apart, and one early treatment scan, obtained during the 2nd week of curative-intent chemoradiation therapy. Regions of interest (ROI), consisting of primary and nodal disease were drawn onto resampled ADC maps. Four 3D texture matrices describing local and regional relationships between voxel intensities in the ROIs were generated. From these, 38 texture metrics and 7 histogram features were calculated for each patient, including the mean and median ADC. Agreement between the two baseline measures was estimated with the intra-class correlation coefficient (ICC). For each metric with an ICC≥0.80, the Wilcoxon signed-rank test was used to test if the difference between the mean of the baselines and the early treatment was non-zero. Results: Texture analysis was implemented on nine patients that had both baselines and early treatment images. Due to baseline agreement, only 9 of the 45 metrics had an ICC ≥0.80, including ADC mean and median. Six of these 9 metrics had a p-value \u3c 0.05. Only 1 of the 9 metrics remained of interest, after applying the Holm correction to the alpha levels: the run length non-uniformity metric (p = 0.004) in the Gray Level Run Length Matrix. Conclusion: The feasibility of texture analysis is dependent on the baseline agreement of each metric, which disqualifies many texture characteristics. However, metrics with high ICC have potential to provide additional quantitative information for the assessment of early treatment changes for HNSCC

Thermal dosimetry characteristics of deep regional heating of non-muscle invasive bladder cancer.

PURPOSE: The aim of this paper is to report thermal dosimetry characteristics of external deep regional pelvic hyperthermia combined with intravesical mitomycin C (MMC) for treating bladder cancer following transurethral resection of bladder tumour, and to use thermal data to evaluate reliability of delivering the prescribed hyperthermia dose to bladder tissue. MATERIALS AND METHODS: A total of 14 patients were treated with MMC and deep regional hyperthermia (BSD-2000, Sigma Ellipse or Sigma 60). The hyperthermia objective was 42° ± 2 °C to bladder tissue for ≥40 min per treatment. Temperatures were monitored with thermistor probes and recorded values were used to calculate thermal dose and evaluate treatment. Anatomical characteristics were examined for possible correlations with heating. RESULTS: Combined with BSD-2000 standard treatment planning and patient feedback, real-time temperature monitoring allowed thermal steering of heat sufficient to attain the prescribed thermal dose to bladder tissue within patient tolerance in 91.6% of treatments. Mean treatment time for bladder tissue \u3e40 °C was 61.9 ± 11.4 min and mean thermal dose was 21.3 ± 16.5 CEM43. Average thermal doses obtained in normal tissues were 1.6 ± 1.2 CEM43 for the rectum and 0.8 ± 1.3 CEM43 in superficial normal tissues. No significant correlation was seen between patient anatomical characteristics and thermal dose achieved in bladder tissue. CONCLUSIONS: This study demonstrates that a hyperthermia prescription of 42° ± 2 °C for 40-60 min can be delivered safely to bladder tissue with external radiofrequency phased array applicators for a typical range of patient sizes. Using the available thermometry and treatment planning, the BSD-2000 hyperthermia system was shown to be an effective method of focusing heat regionally around the bladder with good patient tolerance

Progress in Composite Hydrogels and Scaffolds Enriched with Icariin for Osteochondral Defect Healing

Osteochondral structure reconstruction by tissue engineering, a challenge in regenerative medicine, requires a scaffold that ensures both articular cartilage and subchondral bone remodeling. Functional hydrogels and scaffolds present a strategy for the controlled delivery of signaling molecules (growth factors and therapeutic drugs) and are considered a promising therapeutic approach. Icariin is a pharmacologically-active small molecule of prenylated flavonol glycoside and the main bioactive flavonoid isolated from Epimedium spp. The in vitro and in vivo testing of icariin showed chondrogenic and ostseoinductive effects, comparable to bone morphogenetic proteins, and suggested its use as an alternative to growth factors, representing a low-cost, promising approach for osteochondral regeneration. This paper reviews the complex structure of the osteochondral tissue, underlining the main aspects of osteochondral defects and those specifically occurring in osteoarthritis. The significance of icariin&rsquo;s structure and the extraction methods were emphasized. Studies revealing the valuable chondrogenic and osteogenic effects of icariin for osteochondral restoration were also reviewed. The review highlighted th recent state-of-the-art related to hydrogels and scaffolds enriched with icariin developed as biocompatible materials for osteochondral regeneration strategies