Nitrilotriacetic Acid Improves Plasma Electrolytic Oxidation of Titanium for Biomedical Applications.

Dental implants have become a routine, affordable, and highly reliable technology to replace tooth loss. In this regard, titanium and its alloys are the metals of choice for the manufacture of dental implants because they are chemically inert and biocompatible. However, for special cohorts of patients, there is still a need for improvements, specifically to increase the ability of implants to integrate into the bone and gum tissues and to prevent bacterial infections that can subsequently lead to peri-implantitis and implant failures. Therefore, titanium implants require sophisticated approaches to improve their postoperative healing and long-term stability. Such treatments range from sandblasting to calcium phosphate coating, fluoride application, ultraviolet irradiation, and anodization to increase the bioactivity of the surface. Plasma electrolytic oxidation (PEO) has gained popularity as a method for modifying metal surfaces and delivering the desired mechanical and chemical properties. The outcome of PEO treatment depends on the electrochemical parameters and composition of the bath electrolyte. In this study, we investigated how complexing agents affect the PEO surfaces and found that nitrilotriacetic acid (NTA) can be used to develop efficient PEO protocols. The PEO surfaces generated with NTA in combination with sources of calcium and phosphorus were shown to increase the corrosion resistance of the titanium substrate. They also support cell proliferation and reduce bacterial colonization and, hence, lead to a reduction in failed implants and repeated surgeries. Moreover, NTA is an ecologically favorable chelating agent. These features are necessary for the biomedical industry to be able to contribute to the sustainability of the public healthcare system. Therefore, NTA is proposed to be used as a component of the PEO bath electrolyte to obtain bioactive surface layers with properties desired for next-generation dental implants.

Polycaprolactone-MXene Nanofibrous Scaffolds for Tissue Engineering.

New conductive materials for tissue engineering are needed for the development of regenerative strategies for nervous, muscular, and heart tissues. Polycaprolactone (PCL) is used to obtain biocompatible and biodegradable nanofiber scaffolds by electrospinning. MXenes, a large class of biocompatible 2D nanomaterials, can make polymer scaffolds conductive and hydrophilic. However, an understanding of how their physical properties affect potential biomedical applications is still lacking. We immobilized Ti3C2Tx MXene in several layers on the electrospun PCL membranes and used positron annihilation analysis combined with other techniques to elucidate the defect structure and porosity of nanofiber scaffolds. The polymer base was characterized by the presence of nanopores. The MXene surface layers had abundant vacancies at temperatures of 305-355 K, and a voltage resonance at 8 × 104 Hz with the relaxation time of 6.5 × 106 s was found in the 20-355 K temperature interval. The appearance of a long-lived component of the positron lifetime was observed, which was dependent on the annealing temperature. The study of conductivity of the composite scaffolds in a wide temperature range, including its inductive and capacity components, showed the possibility of the use of MXene-coated PCL membranes as conductive biomaterials. The electronic structure of MXene and the defects formed in its layers were correlated with the biological properties of the scaffolds in vitro and in bacterial adhesion tests. Double and triple MXene coatings formed an appropriate environment for cell attachment and proliferation with mild antibacterial effects. A combination of structural, chemical, electrical, and biological properties of the PCL-MXene composite demonstrated its advantage over the existing conductive scaffolds for tissue engineering.

MXene-Assisted Ablation of Cells with a Pulsed Near-Infrared Laser.

Innovative therapies are urgently needed to combat cancer. Thermal ablation of tumor cells is a promising minimally invasive treatment option. Infrared light can penetrate human tissues and reach superficial malignancies. MXenes are a class of 2D materials that consist of carbides/nitrides of transition metals. The transverse surface plasmons of MXenes allow for efficient light absorption and light-to-heat conversion, making MXenes promising agents for photothermal therapy (PTT). To date, near-infrared (NIR) light lasers have been used in PTT studies explicitly in a continuous mode. We hypothesized that pulsed NIR lasers have certain advantages for the development of tailored PTT treatment targeting tumor cells. The pulsed lasers offer a wide range of controllable parameters, such as power density, duration of pulses, pulse frequency, and so on. Consequently, they can lower the total energy applied and enable the ablation of tumor cells while sparing adjacent healthy tissues. We show for the first time that a pulsed 1064 nm laser could be employed for selective ablation of cells loaded with Ti3C2Tx MXene. We demonstrate both low toxicity and good biocompatibility of this MXene in vitro, as well as a favorable safety profile based on the experiments in vivo. Furthermore, we analyze the interaction of MXene with cells in several cell lines and discuss possible artifacts of commonly used cellular metabolic assays in experiments with MXenes. Overall, these studies provide a basis for the development of efficient and safe protocols for minimally invasive therapies for certain tumors.

MXenes-A New Class of Two-Dimensional Materials: Structure, Properties and Potential Applications.

A new class of two-dimensional nanomaterials, MXenes, which are carbides/nitrides/carbonitrides of transition and refractory metals, has been critically analyzed. Since the synthesis of the first family member in 2011 by Yury Gogotsi and colleagues, MXenes have quickly become attractive for a variety of research fields due to their exceptional properties. Despite the fact that this new family of 2D materials was discovered only about ten years ago, the number of scientific publications related to MXene almost doubles every year. Thus, in 2021 alone, more than 2000 papers are expected to be published, which indicates the relevance and prospects of MXenes. The current paper critically analyzes the structural features, properties, and methods of synthesis of MXenes based on recent available research data. We demonstrate the recent trends of MXene applications in various fields, such as environmental pollution removal and water desalination, energy storage and harvesting, quantum dots, sensors, electrodes, and optical devices. We focus on the most important medical applications: photo-thermal cancer therapy, diagnostics, and antibacterial treatment. The first results on obtaining and studying the structure of high-entropy MXenes are also presented.

Effects of the sources of calcium and phosphorus on the structural and functional properties of ceramic coatings on titanium dental implants produced by plasma electrolytic oxidation.

Plasma Electrolytic Oxidation (PEO) is as a promising technique to modify metal surfaces by application of oxide ceramic coatings with appropriate physical, chemical and biological characteristics. Therefore, objective of this research was to find the simplest settings, yet able to produce relevant bioactive implant surfaces layers on Ti implants by means of PEO. We show that an electrolyte containing potassium dihydrogen phosphate as a source of P and either calcium hydroxide or calcium formate as a source of Ca in combination with a chelating agent, ethylenediamine tetraacetic acid (EDTA), is suitable for PEO to deliver coatings with desired properties. We determined surface morphology, roughness, wettability, chemical and phase composition of titanium after the PEO process. To investigate biocompatibility and bacterial properties of the PEO oxide coatings we used microbial and cell culture tests. The electrolyte based on Ca(OH)2 and EDTA promotes active crystallization of apatites after PEO processing of the Ti implants. The PEO layers can increase electrochemical corrosion resistance. The PEO can be potentially used for development of bioactive surfaces with increased support of eukaryotic cells while inhibiting attachment and growth of bacteria without use of antibacterial agents.

Spinal Reflex Recovery after Dorsal Rhizotomy and Repair with Platelet-Rich Plasma (PRP) Gel Combined with Bioengineered Human Embryonic Stem Cells (hESCs).

Dorsal root rhizotomy (DRZ) is currently considered an untreatable injury, resulting in the loss of sensitive function and usually leading to neuropathic pain. In this context, we recently proposed a new surgical approach to treat DRZ that uses platelet-rich plasma (PRP) gel to restore the spinal reflex. Success was correlated with the reentry of primary afferents into the spinal cord. Here, aiming to enhance previous results, cell therapy with bioengineered human embryonic stem cells (hESCs) to overexpress fibroblast growth factor 2 (FGF2) was combined with PRP. For these experiments, adult female rats were submitted to a unilateral rhizotomy of the lumbar spinal dorsal roots, which was followed by root repair with PRP gel with or without bioengineered hESCs. One week after DRZ, the spinal cords were processed to evaluate changes in the glial response (GFAP and Iba-1) and excitatory synaptic circuits (VGLUT1) by immunofluorescence. Eight weeks postsurgery, the lumbar intumescences were processed for analysis of the repaired microenvironment by transmission electron microscopy. Spinal reflex recovery was evaluated by the electronic Von Frey method for eight weeks. The transcript levels for human FGF2 were over 37-fold higher in the induced hESCs than in the noninduced and the wildtype counterparts. Altogether, the results indicate that the combination of hESCs with PRP gel promoted substantial and prominent axonal regeneration processes after DRZ. Thus, the repair of dorsal roots, if done appropriately, may be considered an approach to regain sensory-motor function after dorsal root axotomy.

Combination of heterologous fibrin sealant and bioengineered human embryonic stem cells to improve regeneration following autogenous sciatic nerve grafting repair.

BACKGROUND: Peripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy. When it is not possible due to a large nerve gap, autologous nerve grafting is used. However, these surgical techniques result in nerve regeneration at highly variable degrees. It is thus very important to seek complementary techniques to improve motor and sensory recovery. One promising approach could be cell therapy. Transplantation therapy with human embryonic stem cells (hESCs) is appealing because these cells are pluripotent and can differentiate into specialized cell types and have self-renewal ability. Therefore, the main objective of this study was to find conditions under which functional recovery is improved after sciatic nerve neurorrhaphy. We assumed that hESC, either alone or in combination with heterologous fibrin sealant scaffold, could be used to support regeneration in a mouse model of sciatic nerve injury and repair via autografting with end-to-end neurorrhaphy. METHODS: Five millimeters of the sciatic nerve of C57BL/6 J mice were transected off and rotated 180 degrees to simulate an injury, and then stumps were sutured. Next, we applied heterologous fibrin sealant and/or human embryonic stem cells genetically altered to overexpress fibroblast growth factor 2 (FGF2) at the site of the injury. The study was designed to include six experimental groups comprising neurorrhaphy (N), neurorrhaphy + heterologous fibrin sealant (N + F), neurorrhaphy + heterologous fibrin sealant + doxycycline (N + F + D), neurorrhaphy + heterologous fibrin sealant + wild-type hESC (N + F + W), neurorrhaphy + heterologous fibrin sealant + hESC off (N + F + T), and neurorrhaphy + heterologous fibrin sealant + hESC on via doxycycline (N + F + D + T). We evaluated the recovery rate using Catwalk and von Frey functional recovery tests, as well as immunohistochemistry analysis. RESULTS: The experiments indicated that sensory function improved when transgenic hESCs were used. The regeneration of sensory fibers indeed led to increased reflexes, upon stimulation of the paw ipsilateral to the lesion, as seen by von-Frey evaluation, which was supported by immunohistochemistry. CONCLUSIONS: Overall, the present data demonstrated that transgenic embryonic stem cells, engineered to overexpress FGF-2 in an inducible fashion, could be employed to support regeneration aiming at the recovery of both motor and sensory functions.

Combination of heterologous fibrin sealant and bioengineered human embryonic stem cells to improve regeneration following autogenous sciatic nerve grafting repair

Background Peripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy. When it is not possible due to a large nerve gap, autologous nerve grafting is used. However, these surgical techniques result in nerve regeneration at highly variable degrees. It is thus very important to seek complementary techniques to improve motor and sensory recovery. One promising approach could be cell therapy. Transplantation therapy with human embryonic stem cells (hESCs) is appealing because these cells are pluripotent and can differentiate into specialized cell types and have self-renewal ability. Therefore, the main objective of this study was to find conditions under which functional recovery is improved after sciatic nerve neurorrhaphy. We assumed that hESC, either alone or in combination with heterologous fibrin sealant scaffold, could be used to support regeneration in a mouse model of sciatic nerve injury and repair via autografting with end-to-end neurorrhaphy. Methods Five millimeters of the sciatic nerve of C57BL/6 J mice were transected off and rotated 180 degrees to simulate an injury, and then stumps were sutured. Next, we applied heterologous fibrin sealant and/or human embryonic stem cells genetically altered to overexpress fibroblast growth factor 2 (FGF2) at the site of the injury. The study was designed to include six experimental groups comprising neurorrhaphy (N), neurorrhaphy + heterologous fibrin sealant (N + F), neurorrhaphy + heterologous fibrin sealant + doxycycline (N + F + D), neurorrhaphy + heterologous fibrin sealant + wild-type hESC (N + F + W), neurorrhaphy + heterologous fibrin sealant + hESC off (N + F +T), and neurorrhaphy + heterologous fibrin sealant + hESC on via doxycycline (N + F + D + T). We evaluated the recovery rate using Catwalk and von Frey functional recovery tests, as well as immunohistochemistry analysis. Results The experiments indicated...

Combination of heterologous fibrin sealant and bioengineered human embryonic stem cells to improve regeneration following autogenous sciatic nerve grafting repair

Peripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy. When it is not possible due to a large nerve gap, autologous nerve grafting is used. However, these surgical techniques result in nerve regeneration at highly variable degrees. It is thus very important to seek complementary techniques to improve motor and sensory recovery. One promising approach could be cell therapy. Transplantation therapy with human embryonic stem cells (hESCs) is appealing because these cells are pluripotent and can differentiate into specialized cell types and have self-renewal ability. Therefore, the main objective of this study was to find conditions under which functional recovery is improved after sciatic nerve neurorrhaphy. We assumed that hESC, either alone or in combination with heterologous fibrin sealant scaffold, could be used to support regeneration in a mouse model of sciatic nerve injury and repair via autografting with end-to-end neurorrhaphy. Methods Five millimeters of the sciatic nerve of C57BL/6 J mice were transected off and rotated 180 degrees to simulate an injury, and then stumps were sutured. Next, we applied heterologous fibrin sealant and/or human embryonic stem cells genetically altered to overexpress fibroblast growth factor 2 (FGF2) at the site of the injury. The study was designed to include six experimental groups comprising neurorrhaphy (N), neurorrhaphy + heterologous fibrin sealant (N + F), neurorrhaphy + heterologous fibrin sealant + doxycycline (N + F + D), neurorrhaphy + heterologous fibrin sealant + wild-type hESC (N + F + W), neurorrhaphy + heterologous fibrin sealant + hESC off (N + F +T), and neurorrhaphy + heterologous fibrin sealant + hESC on via doxycycline (N + F + D + T). We evaluated the recovery rate using Catwalk and von Frey functional recovery tests, as well as immunohistochemistry analysis. Results The experiments indicated that sensory function improved when transgenic hESCs were used. The regeneration of sensory fibers indeed led to increased reflexes, upon stimulation of the paw ipsilateral to the lesion, as seen by von-Frey evaluation, which was supported by immunohistochemistry. Conclusions Overall, the present data demonstrated that transgenic embryonic stem cells, engineered to overexpress FGF-2 in an inducible fashion, could be employed to support regeneration aiming at the recovery of both motor and sensory functions.(AU)

Transgenic human embryonic stem cells overexpressing FGF2 stimulate neuroprotection following spinal cord ventral root avulsion.

Ventral root avulsion (VRA) triggers a strong glial reaction which contributes to neuronal loss, as well as to synaptic detachment. To overcome the degenerative effects of VRA, treatments with neurotrophic factors and stem cells have been proposed. Thus, we investigated neuroprotection elicited by human embryonic stem cells (hESC), modified to overexpress a human fibroblast growth factor 2 (FGF-2), on motoneurons subjected to VRA. Lewis rats were submitted to VRA (L4-L6) and hESC/FGF-2 were applied to the injury site using a fibrin scaffold. The spinal cords were processed to evaluate neuronal survival, synaptic stability, and glial reactivity two weeks post lesion. Then, qRT-PCR was used to assess gene expression of ß2-microglobulin (ß2m), TNFα, IL1ß, IL6 and IL10 in the spinal cord in vivo and FGF2 mRNA levels in hESC in vitro. The results indicate that hESC overexpressing FGF2 significantly rescued avulsed motoneurons, preserving synaptic covering and reducing astroglial reactivity. The cells were also shown to express BDNF and GDNF at the site of injury. Additionally, engraftment of hESC led to a significant reduction in mRNA levels of TNFα at the spinal cord ventral horn, indicating their immunomodulatory properties. Overall, the present data suggest that hESC overexpressing FGF2 are neuroprotective and can shift gene expression towards an anti-inflammatory environment.

Oxidative mechanisms of biological activity of low-intensity radiofrequency radiation.

This review aims to cover experimental data on oxidative effects of low-intensity radiofrequency radiation (RFR) in living cells. Analysis of the currently available peer-reviewed scientific literature reveals molecular effects induced by low-intensity RFR in living cells; this includes significant activation of key pathways generating reactive oxygen species (ROS), activation of peroxidation, oxidative damage of DNA and changes in the activity of antioxidant enzymes. It indicates that among 100 currently available peer-reviewed studies dealing with oxidative effects of low-intensity RFR, in general, 93 confirmed that RFR induces oxidative effects in biological systems. A wide pathogenic potential of the induced ROS and their involvement in cell signaling pathways explains a range of biological/health effects of low-intensity RFR, which include both cancer and non-cancer pathologies. In conclusion, our analysis demonstrates that low-intensity RFR is an expressive oxidative agent for living cells with a high pathogenic potential and that the oxidative stress induced by RFR exposure should be recognized as one of the primary mechanisms of the biological activity of this kind of radiation.

MEK and TGF-beta Inhibition Promotes Reprogramming without the Use of Transcription Factor.

The possibility of replacing the originally discovered and widely used DNA reprogramming transcription factors is stimulating enormous effort to identify more effective compounds that would not alter the genetic information. Here, we describe the generation of induced pluripotent stem cells (iPSc) from head-derived primary culture of mouse embryonic cells using small chemical inhibitors of the MEK and TGF-beta pathways without delivery of exogenous transcription factors. These iPSc express standard pluripotency markers and retain their potential to differentiate into cells of all germ layers. Our data indicate that head-derived embryonic neural cells might have the reprogramming potential while neither the same primary cells cultivated over five passages in vitro nor a cell population derived from adult brain possesses this capacity. Our results reveal the potential for small molecules to functionally replace routinely used transcription factors and lift the veil on molecular regulation controlling pluripotency. The conditions described here could provide a platform upon which other genome non integrative and safer reprogramming processes could be developed. This work also shows novel potential for developing embryonic neural cells.

Nonlinear regression models for determination of nicotinamide adenine dinucleotide content in human embryonic stem cells.

Recent evidence suggests that energy metabolism contributes to molecular mechanisms controlling stem cell identity. For example, human embryonic stem cells (hESCs) receive their metabolic energy mostly via glycolysis rather than mitochondrial oxidative phosphorylation. This suggests a connection of metabolic homeostasis to stemness. Nicotinamide adenine dinucleotide (NAD) is an important cellular redox carrier and a cofactor for various metabolic pathways, including glycolysis. Therefore, accurate determination of NAD cellular levels and dynamics is of growing importance for understanding the physiology of stem cells. Conventional analytic methods for the determination of metabolite levels rely on linear calibration curves. However, in actual practice many two-enzyme cycling assays, such as the assay systems used in this work, display prominently nonlinear behavior. Here we present a diaphorase/lactate dehydrogenase NAD cycling assay optimized for hESCs, together with a mechanism-based, nonlinear regression models for the determination of NAD(+), NADH, and total NAD. We also present experimental data on metabolic homeostasis of hESC under various physiological conditions. We show that NAD(+)/NADH ratio varies considerably with time in culture after routine change of medium, while the total NAD content undergoes relatively minor changes. In addition, we show that the NAD(+)/NADH ratio, as well as the total NAD levels, vary between stem cells and their differentiated counterparts. Importantly, the NAD(+)/NADH ratio was found to be substantially higher in hESC-derived fibroblasts versus hESCs. Overall, our nonlinear mathematical model is applicable to other enzymatic amplification systems.

GSM 900 MHz cellular phone radiation can either stimulate or depress early embryogenesis in Japanese quails depending on the duration of exposure.

PURPOSE: Our study was designed to assess the effects of low intensity radiation of a GSM (Global System for Mobile communication) 900 MHz cellular phone on early embryogenesis in dependence on the duration of exposure. MATERIALS AND METHODS: Embryos of Japanese Quails were exposed in ovo to GSM 900 MHz cellular phone radiation during initial 38 h of brooding or alternatively during 158 h (120 h before brooding plus initial 38 h of brooding) discontinuously with 48 sec ON (average power density 0.25 µW/cm(2), specific absorption rate 3 µW/kg) followed by 12 sec OFF intervals. A number of differentiated somites were assessed microscopically. Possible DNA damage evoked by irradiation was assessed by an alkaline comet assay. RESULTS: Exposure to radiation from a GSM 900 MHz cellular phone led to a significantly altered number of differentiated somites. In embryos irradiated during 38 h the number of differentiated somites increased (p < 0.001), while in embryos irradiated during 158 h this number decreased (p < 0.05). The lower duration of exposure led to a significant (p < 0.001) decrease in a level of DNA strand breaks in cells of 38-h embryos, while the higher duration of exposure resulted in a significant (p < 0.001) increase in DNA damage as compared to the control. CONCLUSION: Effects of GSM 900 MHz cellular phone radiation on early embryogenesis can be either stimulating or deleterious depending on the duration of exposure.

Adaptation to robust monolayer expansion produces human pluripotent stem cells with improved viability.

The generation of human pluripotent stem cells (hPSCs) of sufficient quantity and quality remains a major challenge for biomedical application. Here we present an efficient feeder-free, high-density monolayer system in which hPSCs become SSEA-3-high and gradually more viable than their feeder-dependent counterparts without changes attributed to culture adaptation. As a consequence, monolayer hPSCs possess advantages over their counterparts in embryoid body development, teratoma formation, freezing as a single-cell suspension, and colony-forming efficiency. Importantly, this monolayer culture system is reversible, preserving the competence of hPSCs to gradually reacquire features of colony growth, if necessary. Therefore, the monolayer culture system is highly suitable for long-term, large-scale propagation of hPSCs, which is necessary in drug development and pluripotent stem cell-based therapies.

GSM 900 MHz microwave radiation affects embryo development of Japanese quails.

A wide range of non thermal biological effects of microwave radiation (MW) was revealed during the last decades. A number of reports showed evident hazardous effects of MW on embryo development in chicken. In this study, we aimed at elucidating the effects of MW emitted by a commercial model of GSM 900 MHz cell phone on embryo development in quails (Coturnix coturnix japonica) during both short and prolonged exposure. For that, fresh fertilized eggs were irradiated during the first 38 h or 14 days of incubation by a cell phone in "connecting" mode activated continuously through a computer system. Maximum intensity of incident radiation on the egg's surface was 0.2 µW/cm2.The irradiation led to a significant (p<0.001) increase in numbers of differentiated somites in 38-hour exposed embryos and to a significant (p<0.05) increase in total survival of embryos from exposed eggs after 14 days exposure. We hypothesized that observed facilitating effect was due to enhancement of metabolism in exposed embryos provoked via peroxidation mechanisms. Indeed, a level of thiobarbituric acid (TBA) reactive substances was significantly (p<0.05-0.001) higher in brains and livers of hatchlings from exposed embryos. Thus, observed effects of radiation from commercial GSM 900 MHz cell phone on developing quail embryos signify a possibility for non-thermal impact of MW on embryogenesis. We suggest that the facilitating effect of low doses of irradiation on embryo development can be explained by a hormesis effect induced by reactive oxygen species (ROS). Future studies need to be done to clarify this assumption.

Trends and co-trends of prostate-specific antigen and body mass index in a screened population.

OBJECTIVES: This report investigated whether annual changes in body mass index (BMI) are associated with the opposite changes in prostate-specific antigen (PSA). Previous studies have confirmed lower PSA levels among men with higher BMI. METHODS: Normal linear mixed models were used to characterize annual PSA, BMI and the ratio of PSA to BMI profiles for 2641 men undergoing prostate cancer screening for up to 8 years as part of a San Antonio screening study. RESULTS: Among the 1898 participants (71.9%) who never received a prostate biopsy during the study and the 585 participants (22.1%) who had one or more biopsies, all negative for prostate cancer, BMI was higher for Hispanics than other racial groups, lower for older men at study entry, and increased every year during the study; and PSA and PSA/BMI ratios were higher for older men at study entry and increased each year on study (all P values<.05). Among the 158 men (6.0%) eventually diagnosed with prostate cancer, no trends in BMI were statistically significant, but PSA and PSA/BMI ratios were higher on average for older men at study entry and increased each year on study (both P values<.05). Correlations between BMI and PSA changes per year were negative but not statistically significantly different from zero. CONCLUSIONS: The individual man scrutinizing his PSA and weight year to year can expect a slight annual increase in both, but changes in PSA from one year to the next cannot be attributed to weight gain or loss.

Sodium butyrate induces cellular senescence in neuroblastoma and prostate cancer cells.

Cellular senescence leads to an irreversible block of cellular division capacity both in cell culture and in vivo. The induction of an irreversible cell cycle arrest is very useful for treatment of cancer. Histone deacetylases (HDACs) are considered as therapeutic targets to treat cancer patients. HDAC inhibitors repress cancer growth and are used in various clinical trials. Here, we analyzed whether sodium butyrate (NaBu), an inhibitor of class I and II HDACs, induces cellular senescence in neuroblastoma and prostate cancer (PCa) including an androgen-dependent as well as an androgen-independent human PCa cell line. We found that the HDAC inhibitors NaBu and valproic acid (VPA) induce cellular senescence in tumor cells. Interestingly, also an inhibitor of SIRT1, a class HDAC III, induces cellular senescence. Both neuroblastoma and human prostate cancer cell lines express senescence markers, such as the Senescence Associated-ß-galactosidase (SA-ß-Gal) and Senescence Associated Heterochromatin Foci (SAHF). Furthermore, NaBu down-regulates the proto-oncogenes c-Myc, Cyclin D1 and E2F1 mRNA levels. The mRNA level of the cell cycle inhibitor p16 remains unchanged whereas that of the tumor suppressor p21 is strongly up-regulated. Interestingly, NaBu treatment robustly increases reactive oxygen species (ROS) levels. These results indicate an epigenetic regulation and an association of HDAC inhibition and ROS production with cellular senescence. The data underline that tumor cells can be driven towards cellular senescence by HDAC inhibitors, which may further arise as a potent possibility for tumor suppression.

N(epsilon)-thioacetyl-lysine-containing tri-, tetra-, and pentapeptides as SIRT1 and SIRT2 inhibitors.

N()-Thioacetyl-lysine-containing tri-, tetra-, and pentapeptides, based on the alpha-tubulin and p53 protein sequences, were studied as SIRT1 and SIRT2 inhibitors. The potency of the pentapeptides depended on the selection of the side chains. The removal of N- and C-terminal residues of the pentapeptides yielded tripeptides with retained SIRT1 inhibitory activity but decreased SIRT2 inhibitory activity. The most potent SIRT1 inhibitors were equipotent with the reference compound (6-chloro-2,3,4,9-tetrahydro-1H-carbazole-1-carboxamide) with the IC(50) values of 180-330 nM.

Characterization of the binding properties of SIRT2 inhibitors with a N-(3-phenylpropenoyl)-glycine tryptamide backbone.

SIRT2 inhibitors with a N-(3-phenylpropenoyl)-glycine tryptamide backbone were studied. This backbone has been developed in our group, and it is derived from a compound originally found by virtual screening. In addition, compounds with a smaller 3-phenylpropenoic acid tryptamide backbone were also included in the study. Binding modes for the new compounds and the previously reported compounds were analyzed with molecular modelling methods. The approach, which included a combination of molecular dynamics, molecular docking and cluster analysis, showed that certain docking poses were favourable despite the conformational variation in the target protein. The N-(3-phenylpropenoyl)-glycine tryptamide backbone is also a good backbone for SIRT2 inhibitors, and the series of compounds includes several potent SIRT2 inhibitors.