Enhancing Temozolomide (TMZ) chemosensitivity using CRISPR-dCas9-mediated downregulation of O6-methylguanine DNA methyltransferase (MGMT).

PURPOSE: Glioblastoma (GBM) stands out as the most prevalent and aggressive intracranial tumor, notorious for its poor prognosis. The current standard-of-care for GBM patients involves surgical resection followed by radiotherapy, combined with concurrent and adjuvant chemotherapy using Temozolomide (TMZ). The effectiveness of TMZ primarily relies on the activity of O6-methylguanine DNA methyltransferase (MGMT), which removes alkyl adducts from the O6 position of guanine at the DNA level, thereby counteracting the toxic effects of TMZ. METHOD: In this study, we employed fusions of catalytically-inactive Cas9 (dCas9) to DNA methyltransferases (dCas9-DNMT3A) to selectively downregulation MGMT transcription by inducing methylation at MGMT promoter and K-M enhancer. RESULT: Our findings demonstrate a significant reduction in MGMT expression, leading to intensified TMZ sensitivity in the HEK293T cell line. CONCLUSION: This study serves as a proof of concept for the utilization of CRISPR-based gene suppression to overcome TMZ resistance and enhance the lethal effect of TMZ in glioblastoma tumor cells.

AR13 peptide-conjugated liposomes improve the antitumor efficacy of doxorubicin in mice bearing C26 colon carcinoma; in silico, in vitro, and in vivo study.

Currently, liposomes have emerged as efficient and safer nano-carriers for targeted therapy in different cancers. This work aimed to employ PEGylated liposomal doxorubicin (Doxil®/PLD), modified with AR13 peptide, to target Muc1 on the surface of colon cancerous cells. We performed molecular docking and simulation studies (using Gromacs package) of AR13 peptide against Muc1 to analyze and visualize the peptide-Muc1 binding combination. For in vitro analysis, the AR13 peptide was post-inserted into Doxil® and verified by TLC, 1H NMR, and HPLC techniques. The zeta potential, TEM, release, cell uptake, competition assay, and cytotoxicity studies were performed. In vivo antitumor activities and survival analysis on mice bearing C26 colon carcinoma were studied. Results showed that after 100 ns simulation, a stable complex between AR13 and Muc1 formed, and molecular dynamics analysis confirmed this interaction. In vitro analysis demonstrated significant enhancement of cellular binding and cell uptake. The results of in vivo study on BALB/c mice bearing C26 colon carcinoma, revealed an extended survival time to 44 days and higher tumor growth inhibition compared to Doxil®. Thus, the AR13 peptide could be explored as a potent ligand for Muc1, improving therapeutic antitumor efficiency in colon cancer cells.

Preparation of chitosan nanoparticle containing recombinant CD44v antigen and evaluation of its immunization capacity against breast cancer in BALB/c mice.

OBJECTIVE(S): Breast tumors show heterogeneity containing cancer stem cells as a small subpopulation of a tumor mass. CD44 as a cancer stem cells antigen is abnormally expressed by carcinomas of epithelial origin. Also, overexpression of CD44 variable isoforms (CD44v) is associated with malignancy in breast cancer. In the present research, our objective was to evaluate the immunogenicity of prepared nanoparticles containing a novel recombinant CD44v (rCD44v) protein in the mouse model. MATERIALS AND METHODS: CD44 gene was expressed in E. coli BL21 DE3 using the pET28a-CD44 vector. The expressed rCD44v protein was purified, encapsulated into the chitosan nanoparticles, and administered to BALB/c mice. ELISA was used to evaluate the immunoglobulin levels of immunized animals. For challenge experiment, 2 × 106 4T1-CD44 tumor cells were injected subcutaneously in mice, and tumor size, necrosis, and metastases were measured. Finally, cell proliferation assay, cytokines assay, and neutralization assay of the mouse anti-rCD44v on the human breast cancer cell line were examined. RESULTS: The measured size of chitosan-rCD44v nanoparticles was 146.5 nm. Recombinant CD44v encapsulated by chitosan nanoparticles increases immunological responses via the adjuvant nature of chitosan nanoparticles. In the immunized mice, IgG and IgA titers were significantly increased. Tumor growth in injection and nano-injection test groups compared with the mice control groups displayed a significant reduction (P < 0.05). A high amount of splenocytes secreting IFNγ and IL-17 was seen in immunized mice with rCD44v (P < 0.05). Furthermore, a smaller size of lung metastases compared to the control mice groups was detected. CONCLUSION: The encapsulated rCD44v within the chitosan nanoparticles induced a significant immune response in mice and can establish significant protection against breast cancer. Therefore, it can be considered a vaccine candidate for breast cancer therapeutic modalities.

Co-administration of curcumin with other phytochemicals improves anticancer activity by regulating multiple molecular targets.

Natural plant phytochemicals are effective against different types of diseases, including cancer. Curcumin, a powerful herbal polyphenol, exerts inhibitory effects on cancer cell proliferation, angiogenesis, invasion, and metastasis through interaction with different molecular targets. However, the clinical use of curcumin is limited due to poor solubility in water and metabolism in the liver and intestine. The synergistic effects of curcumin with some phytochemicals such as resveratrol, quercetin, epigallocatechin-3-gallate, and piperine can improve its clinical efficacy in cancer treatment. The present review specifically focuses on anticancer mechanisms related to the co-administration of curcumin with other phytochemicals, including resveratrol, quercetin, epigallocatechin-3-gallate, and piperine. According to the molecular evidence, the phytochemical combinations exert synergistic effects on suppressing cell proliferation, reducing cellular invasion, and inducing apoptosis and cell cycle arrest. This review also emphasizes the significance of the co-delivery vehicles-based nanoparticles of such bioactive phytochemicals that could improve their bioavailability and reduce their systemic dose. Further high-quality studies are needed to firmly establish the clinical efficacy of the phytochemical combinations.

The effect of angiotensin receptor blockers and angiotensin-converting enzyme inhibitors on progression of gastric cancer: systematic review and meta-analysis.

Angiotensin receptor blockers (ARB), as well as angiotensin-converting enzyme inhibitors (ACEI), are mostly used as therapy for hypertension and cardiovascular disease. However, they can increase the risk of cancer progression including gastric cancer. Here we aimed to analyze the assessment between ARB and ACEI on the progression of gastric cancer. Cochrane Library, PubMed and EMBASE were searched for articles and abstracts describing ARBs, ACEIs, and incidence of gastric cancer. Risk ratio, hazard ratio and 95% confidence interval (CI) were extracted from each outcome by using a random-effects model. Six studies met our inclusion criteria. These results demonstrated that there is a significant association between ARB with gastric cancer progression (risk ratio = 0.63; 95% CI, 0.5-0.7; P = 0.00; I 2 = 27.299; df (Q) = 2; Q-value = 2.75). However, there was not any link between ACEIs and gastric cancer development (risk ratio = 1.1; 95% CI, 0.92-1.31; P = 0.26; I 2 = 0.00; df (Q) = 3; Q-value = 1.26). All these findings indicated that using the ARBs has raised the progression of gastric cancer in these patients.

The expression of long non-coding RNA LINC01389, LINC00365, RP11-138J23.1, and RP11-354K4.2 in gastric cancer and their impacts on EMT.

BACKGROUND: Epithelial cancers acquire the epithelial to mesenchymal transition (EMT), which leads tumor cells to invade and metastasize to adjacent and distant tissues. The mechanisms involved in EMT phenotype are controlled by numerous markers as well as signalling pathways. Recently, long non-coding RNAs (lncRNAs) were introduced that play the regulatory role in EMT via crosstalk with EMT-related transcription factors and signalling pathways. The present study aimed to investigate the expression of four lncRNAs in human GC and elucidate their probable role in EMT procedure and the pathogenesis of gastric cancer (GC). METHODS: The expression profile of lncRNAs (LINC01389, LINC00365, RP11-138J23.1, and RP11-354K4.2) and mRNAs (TWIST1, MMP13, MAML1, CD44s, and SALL4) between eighty-three GC and adjacent non-cancerous tissues were assessed by quantitative real-time PCR. RESULTS: The significant downregulation of LINC00365 (66.3%) and RP11-354K4.2 (62.7%) were observed in GC samples; while the upregulation of LINC01389, RP11-138J23.1, TWIST1, MMP13, MAML1, CD44s, and SALL4 were found in 67.5%, 45.8%, 56.6%, 44.6%, 59%, 55.4%, and 62.7% tumors samples at the mRNA level, respectively. Dysregulation of these lncRNAs and EMT-related markers was significantly related to each other in a variety of clinicopathological features of patients (P < 0.05), indicating positive correlations between LINC01389, LINC00365, RP11-138J23.1, and RP11-354K4.2 with EMT status in GC. CONCLUSION: These EMT-regulating lncRNAs may play a key role in transforming gastric epithelial to mesenchymal phenotype and can be novel therapeutic targets for GC. Our results highlight the importance of discovering new lncRNAs involved in gastric carcinogenesis. Detailed molecular mechanisms of these noncoding-coding markers in GC are urgently required.

Evaluation of the in vivo antihypertensive effect and antioxidant activity of HL-7 and HL-10 peptide in mice.

BACKGROUND: The tendency to use bioactive peptides has increased in recent decades, and research would be essential for recognizing the therapeutic effects of peptides present in animals or food resource. In this study, the in vivo antioxidant and antihypertensive properties of peptides HL-7 with the sequence of YLYELR and HL-10 with the sequence of AFPYYGHHLG were identified from scorpion venom of H. lepturus were evaluated. METHODS AND RESULTS: To study the in vivo effects of peptides, D-galactose-induced and DOCA salt-induced mice models were used. The results of the antioxidant assay for both peptides showed that the activity of serum and liver catalase (CAT), as well as superoxide dismutase (SOD) enzymes, was significantly decreased in the D-galactose-induced group (NC), while MDA levels were increased in serum and the liver tissue samples (p < 0.01). Compared with the D-galactose-induced mice, the peptide treated mice group had a higher activity of antioxidant enzymes namely CAT and SOD, as well as a lower lipid peroxidation level. Also, the results of antihypertensive activity for both peptides showed that systolic blood pressure (SBP) and diastolic blood pressure (DBP) of the mice treated with the HL-7 and HL-10 peptides were significantly reduced in a dose-dependent manner (p < 0.01). The administration of the HL-7 peptide at doses of 2 mg/kg BW (LP1), 5 mg/kg BW (-IP1) and 15 mg/kg BW (HP1) significantly diminished the mean arterial blood pressure (MAP) by 11 mmHg, 31 mmHg and 40.47 mmHg, respectively. Accordingly, treatment of mice with the HL-10 peptide at doses of 2 mg/kg BW (LP2), 5 mg/kg BW (IP2) and 15 mg/kg BW (HP2) considerably lowered the MAP by 8 mmHg, 18.3 mmHg and 21.93 mmHg, respectively. CONCLUSION: Our findings suggest that both the HL-7 and HL-10 peptides could be potentially utilized as antihypertensive and antioxidant components.

Biochemical characterization of an alkaline surfactant-stable keratinase from a new keratinase producer, Bacillus zhangzhouensis.

A new keratinase producer, Bacillus sp. BK111, isolated from a poultry feather was identified as Bacillus zhangzhouensis, which is the first report for its keratinolytic activity. The keratinase production was optimized, followed by the enzyme purification and characterization using biochemical assays. A 2.34-fold increase was observed in the enzyme production under optimized conditions. The enzyme was characterized as a serine protease with 42 kDa molecular weight, stable in a wide range of temperature and pH with maximum keratinolytic activity at 60 °C and pH 9.5. The enzyme had a wide range of different substrates with the best performance on the feather meal substrate. Metal ions of Ca2+, K+, Na+ and Mn2+ enhanced the enzyme activity. The enzyme showed a great deal of stability in the presence of ethanol, methanol, acetone, 2-propanol, dimethyl sulfoxide, Tween-80 and Triton X-100. Dithiothreitol (DTT), as a reducing agent, caused a twofold increase in keratinolytic activity. The half-life of the enzyme at optimum temperature was calculated to be 125 min and the ratio of keratinolytic:caseinolytic for the enzyme was 0.8. Our results showed the remarkable features of the enzyme that make it suitable for biotechnological usages.

Purification and Biochemical Characterization of Alkalophilic Cellulase from the Symbiotic Bacillus subtilis BC1 of the Leopard Moth, Zeuzera pyrina (L.) (Lepidoptera: Cossidae).

In the current study, an extracellular cellulase belonging to symbiotic Bacillus subtilis Bc1 of the leopard moth is purified and characterized. The molecular mass of enzyme was 47.8 kDa using SDS-PAGE. The purified enzyme had optimum activity in temperature and pH around 60 °C and 8, respectively. The purified cellulase was introduced as a stable enzyme in a wide variety of temperature (20-80 °C) and pH (4-10) and remained active to more than 74% at 80 °C for 1 h. Moreover, the cellulase extremely was stabled in the presence of metal ions and organic solvents and its activity was increased by acetone (20% v/v), CaCl2 and CoCl2 and inhibited by MnCl2 and NiCl2. The values of enzyme's Km and Vmax were found to be 1.243 mg/mL and 271.3 µg/mL/min, respectively. The purified cellulase hydrolyzed cellulose, avicel and carboxymethyl cellulose (CMC) and the final product of CMC hydrolysis was cellobiose using thin-layer chromatography analysis. Consequently, owing to exo/endoglucanase activity and organic solvent, temperature and pH stability of the purified cellulase belong to B. subtilis BC1, it can be properly employed for various industrial purposes.

Comparative study of different forms of Jellein antimicrobial peptide on Leishmania parasite.

Typically, antimicrobial peptides (AMPs) are short positive charged peptides serving a key role in innate immunity as well as antimicrobial activity. Discovering novel therapeutic agents is considered as an undeniable demand due to increasing microbial species with antibiotic resistance. In this direction, the unique ability of AMPs to modulate immune responses highlighted them as novel drug candidates in the field of microbiology. Patients affected by leishmaniasis; a neglected tropical disease, confront serious problems for their treatment including resistance to common drugs as well as toxicity and high cost of therapy. So, there is a need for development of new drug candidates to control the diseases. Jellein, a peptide derived from royal jelly of honeybee has been shown to have promising effect against several bacterial and fungal species. In current study, anti-leishmanial effect of Jellein and its lauric acid conjugated form was investigated against two forms of Leishmania major (L. major) parasite. Moreover, cytotoxic effect of these peptides was studied in THP1 cell line and human Red Blood Cells (RBCs). Furthermore, the mechanism of action of peptides on L. major promastigotes was assessed through different methods. The results demonstrated that, conjugation of lauric acid to Jellein not only had no effect on the elevation of antimicrobial activity but also halted it completely. Moreover, Jellein caused a limitation in the number of L. major promastigotes by pore formation as well as changing the membrane potential rather than induction of apoptosis or activation of caspases.

Laccase mediator system obtained from a marine spore exhibits decolorization potential in harsh environmental conditions.

Laccases play a significant role in remedying dye pollutants. Most of these enzymes are originated from terrestrial fungi and bacteria, thus they are not proper to be used in the environments with neutral/alkaline pH, or they may require laborious extraction/purification steps. These limitations can be solved using marine spore laccases through high stability and easy to use application. In the current study, laccase activity of the marine spore -forming Bacillus sp. KC2 was measured according to the guaiacol and syringaldazine oxidation. Abiotic stresses like pH of 6, temperature of 37 °C and 0.3 mM CuSO4 (in comparison with optimal sporulation conditions: pH of 8, temperature of 20 °C and 0.0 mM CuSO4) enhanced laccase formation in sporal coat. Maximum activity of enzyme was observed at 50 °C and pH 7, which did not change in the alkaline pH and temperature range of 20-70 °C. Results indicated ions, inhibitors and solvent stability of the enzyme and its activity were stimulated by Co2+, Mn2+, PMSF, acetone, acetonitrile, ethanol, and methanol. The spore laccase could decolorize synthetic dyes from various chemical groups including azo (acid orange, amaranth, trypan blue, congo red, and amido black), indigo (indigo carmine), thiazine (methylene blue, and toluidine blue), and triarylmethane (malachite green) with ABTS/syringaldazine mediators after 5 h. Degradation products were not toxic against Sorghum vulgare and Artemia salina model organisms. The enzyme mediator system showed high potentials for dye bioremediation over a wide range of harsh conditions.

Antioxidant and angiotensin-converting enzyme (ACE) inhibitory activity of thymosin alpha-1 (Thα1) peptide.

In this research, the antioxidant property of thymosin alpha-1 (Thα1) peptide was investigated through various antioxidant methods. Thα1 showed 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (IC50 = 20 µM) and its 2,2-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) scavenging reached 45.33% at 80 µM (IC50 = 85 µM). In addition, hydroxyl and superoxide radical scavenging of Thα1 peptide exhibited a concentration-depended manner. The IC50 values of hydroxyl and superoxide radical scavenging were estimated to be 82 µM and 20 µM, respectively. The effect of Thα1 on eliminating superoxide radicals was higher (62.23%) than other antioxidant assays. Moreover, the antioxidant activity of Thα1 peptide was evaluated by measuring cellular reactive oxygen species (ROS). Results indicated that Thα1 decreased the generation of ROS level in 1321 N1 human neural asterocytoma cells. The inhibitory effect of Thα1 on angiotensin-converting enzyme (ACE) was determined. The kinetic parameters (Km and Vmax) and the inhibition pattern were examined. Based on the Lineweaver-Burk plot, Thα1 displayed a mixed inhibition pattern. The IC50 and Ki values of Thα1 were 0.8 µM and 3.33 µM, respectively. Molecular modeling suggested that Thα1 binds to ACE-domains with higher affinity binding to N-domain with the binding energy of -22.87 kcal/mol. Molecular docking indicated that Thα1 interacted with ACE enzyme (N- and C-domains) due to electrostatic, hydrophobic, and hydrogen forces. Our findings suggested that Thα1 possess a multifunctional peptide with dual antioxidant and ACE-inhibitory properties. Further researches are needed to investigate the antioxidant and anti-hypertensive effect of Thα1 both in vitro and in vivo.

Thymosin alpha-1; a natural peptide inhibits cellular proliferation, cell migration, the level of reactive oxygen species and promotes the activity of antioxidant enzymes in human lung epithelial adenocarcinoma cell line (A549).

This research was conducted to investigate the biochemical effects of thymosin alpha-1 using human lung cancer cells (A549). The A549 cells were treated with different concentrations of Thα1 for 24 h and the growth, inhibition of cells was determined. Thα1 revealed anti-proliferative effect at 24 and 48 µg/ml after 24 h. Furthermore, it indicated antioxidant properties by significantly enhancing the activity of catalase (12 µg/ml), superoxide dismutase (6 and 12 µg/ml), and glutathione peroxidase (3, 6 and 12 µg/ml) and reducing the production of cellular ROS. Our results showed that Thα1 inhibits the migration of A549 cells in a concentration-dependent manner after 24 and 48 h. Moreover, the effect of Thα1 on apoptosis was investigated by Hoechst 33342 staining and cell cycle analysis. Results demonstrated no significant effect on the induction of apoptosis in A549 cells. In conclusion, our results showed the antioxidant properties of Thα1 on A549 cancer cells.

Biodegradation of phenol by cold-tolerant bacteria isolated from alpine soils of Binaloud Mountains in Iran.

Degradation of phenol is considered to be a challenge because of harsh environments in cold regions and ground waters. Molecular characterization of phenol degrading bacteria was investigated to gain an insight into the biodegradation in cold areas. The psychrotolerant and psychrophiles bacteria were isolated from alpine soils in the northeast of Iran. These strains belonged to Pseudomonas sp., Stenotrophomonas spp. and Shinella spp. based on analysis of the 16S rRNA gene. These strains were capable of the complete phenol degradation at a concentration of 200 mg L-1 at 20 °C. Moreover, the strains could degrade phenol at a concentration of 400 and 600 mg L-1 at a higher time. Effects of environmental factors were studied using one factor at a time (OFAT) approach for Pseudomonas sp.ATR208. When the bacterium was grown in a liquid medium with 600 mg L-1 of concentration supplemented with optimum carbon and nitrogen sources, more than 99% of phenol removal was obtained at 20 °C and 24 h. Therefore, the present study indicated the potential of the local cold tolerant bacteria in the phenol bioremediation.

The inhibitory activity of HL-7 and HL-10 peptide from scorpion venom (Hemiscorpius lepturus) on angiotensin converting enzyme: Kinetic and docking study.

The hypertension is one of the highest risk factors for stroke, myocardial infarction, vascular disease and chronic kidney disease. Angiotensin converting enzyme (ACE) has an important role in the physiological regulation of cardiovascular system. ACE inhibition is a key purpose for hypertension treatment. In this study, two peptides named HL-7 with the sequence of YLYELAR (MW: 927.07Da) and HL-10 with the sequence of AFPYYGHHLG (MW: 1161.28Da) were identified from scorpion venom of H. lepturus. The inhibitory activity of HL-7 and HL-10 was examined on rabbit ACE. The inhibition mechanisms were assayed by kinetic and docking studies. The IC50 values for ACE inhibition of HL-7 and HL-10 were 9.37µM and 17.22µM, respectively. Lineweaver-Burk plots showed that two peptides inhibited rabbit ACE with competitive manner. The molecular docking conformed experimental results and showed that the two peptides interacted with N-domain and C-domain active sites. Also, docking study revealed that the two peptides can form hydrogen and hydrophobic bonds at their binding sites. Both peptides had higher affinity to N-domain. Our results showed that HL-7 exhibited more strong interactions with amino acids at active site. It seems that HL-10 peptide could occupy more space, thereby inhibiting the substrate entrance to active site.

GL-9 peptide regulates gene expression of CD44 cancer marker and pro-inflammatory cytokine TNF-α in human lung epithelial adenocarcinoma cell line (A549).

In this study, we examined cytotoxic effect of GL-9 peptide on A459 cell line through studying the changes in TNF-α and CD44 gene expression and ROS production. Real-time PCR analysis showed that the treated A549 cells highly over expressed TNF-α, which was associated with a significant reduction of CD44 gene expression levels (p  <  0.05). ROS production rate was measured through the usage of DCFH-DA primer. Results demonstrated that GL-9 peptide could also induce cell death via ROS production. The effect of GL-9 peptide on human erythrocytes and leukocytes was analyzed. GL-9 peptide showed no significant toxic effect on human blood cells. Our results suggested that the GL-9 peptide as a potent natural agent could modulate gene expression of cancer cell markers.

An identified antioxidant peptide obtained from ostrich (Struthio camelus) egg white protein hydrolysate shows wound healing properties.

CONTEXT: Ostrich (Struthio camelus) egg possesses a high amount of food proteins and thus plays an important role in nutrition. OBJECTIVE: Ostrich egg white proteins were hydrolyzed with pepsin and pancreatin to examine its antioxidant properties and further characterized the most active peptide. MATERIALS AND METHODS: Ostrich egg white protein hydrolysate (OEWPH) was fractionized using reversed phase high-pressure liquid chromatography (HPLC). The antioxidant activity of OEWPH and its HPLC fraction were investigated based on their scavenging capacity1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, 2,2'-azinobis (3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS), superoxide ([Formula: see text]), hydroxyl (OH(•-)) radicals, and Cu(+2) chelating. In a wound healing assay, paravertebral excision (1 cm diameter) was made on the skin and the percentage of wound closure was measured at defined intervals (0, 3, 7, and 14 d). RESULTS: A potent antioxidant peptide named DG-10 with the sequence DAESLSRLLG (MW: 1060.18 ± 0.5 Da) was identified from OEWPH. The peptide DG-10 showed DPPH (IC50 = 0.0085 mg/ml), ABTS(•+) (IC50 = 0.56 mg/ml), superoxide (IC50 = 0.36 mg/ml), and hydroxyl (IC50 = 0.4 mg/ml) radical scavenger and copper chelating activity (IC50 = 0.28 mg/ml). In vitro cultured HFLF-pI 5, the cell model, also revealed that DG-10 could protect HFLF-pI 5 cells against H2O2-treated necrosis. Ointment composed of DG-10 peptide exhibited wound-healing properties on adult rats (Wistar strain). The percentage of wound closure in peptide-treated group was 98% by day 14. DISCUSSION AND CONCLUSION: Our results suggested that DG-10 is a natural agent obtained from ostrich egg possessing considerable antioxidant and wound-healing properties.

Antioxidant properties of a human neuropeptide and its protective effect on free radical-induced DNA damage.

Human catestatin CgA352-372 (SL21) is an endogenous neuropeptide with multiple biological functions. The present study aimed to evaluate the antioxidant, antibacterial, cytotoxic, and DNA damage protective effects of SL21 neuropeptide. SL21 neuropeptide generated from the C-terminus of chromogranin A (CgA) was synthesized by solid-phase method. Synthetic peptide was subjected to various in vitro antioxidant assays including the scavenging of 1,1-diphenyl-2-pycryl-hydrazyl (DPPH), 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS(·+) ), and hydroxyl free radicals, metal ion chelation, inhibition of lipid peroxidation, and reducing power. Moreover, protective effect of SL21 on H2 O2 -induced DNA damage was analyzed using pTZ57/RT plasmid. Methylthiazoltetrazolium assay was also performed to study the cytotoxic effect of SL21 neuropeptide on human peripheral blood mononuclear cells. Furthermore, antibacterial and hemolysis assays were conducted. The results demonstrated high activities of SL21 in scavenging free radicals (DPPH, ABTS(·+) , and hydroxyl), chelating of Cu(2+) /Fe(2+) metal ions, reducing power, and inhibition of lipid peroxidation in a concentration-dependent manner. SL21 neuropeptide revealed a protective effect on DNA damage caused by hydroxyl radicals. Interestingly, the peptide exhibited no significant cytotoxicity towards peripheral blood mononuclear cells. Furthermore, SL21 peptide displayed antimicrobial activity against Staphylococcus aureus and Pseudomonas aeruginosa without any hemolytic activity on human red blood cells. Conclusively, the present study established SL21 (catestatin) as a novel antioxidative peptide that could further be investigated for its potential use as a pharmaceutical agent.

An overview of Brevinin superfamily: structure, function and clinical perspectives.

Antimicrobial peptides are the backbone of first-line defense against various microorganisms in the animal kingdom. Thus, not surprisingly, they are gaining attention in the science and medical fields as a rich repository of new pro-drugs. Below, we focus our attention on the Brevinin family of anuran peptides. While most of them show strong antibacterial activities, some, e.g. Brevinin-2R, appear to be promising anticancer molecules, exhibiting better a therapeutic window than widely-use anticancer drugs like doxorubicin. We briefly introduce the field, followed by highlighting the promising therapeutic properties of Brevinins. Next, we provide information about the cloning and phylogenetic aspects of Brevinin genes. In the final paragraphs of this chapter, we discuss possible large-scale production methods of Brevinins, giving examples of some systems that are already in use. Towards the end, we discuss various means of modification of biologic properties of Brevinins, either by chemical modifications or by amino acid substitution and sequence rearrangements. In this context, also other unique properties of Brevinins are briefly mentioned. Finally, we discuss the future of the Brevinin field, particularly highlighting yet to be answered biologic questions, like for example presumed anti-viral and antitumor activities of Brevinin family members.

Development of a highly porous bioscaffold by the combination of bubble entrapping and freezing-thawing techniques to fabricate hyaluronic acid/gelatin tri-layer wound dressing.

The electrospun nanofibers and porous scaffolds hold great promise in regenerative medicine. A novel nanofiber-hydrogel­silicone tri-layer wound dressing has been designed and fabricated to address the limitations of each platform. The bottom nanofiber layer with a 110 mm diameter meets the wound surface and regulates cell attachment and migration. The middle hydrogel layer was fabricated through the optimization of chemical crosslink formation and freezing-thawing cycles (physical crosslink). The fabricated hydrogel with interconnected porous structure has optimized properties (gel fraction (89.45 %) and porosity (80 %)) for wound dressing application. The silicone layer on the outer surface was designed to fix the wound dressing on the skin and prevent the penetration of pathogens. The scanning electron microscope micrograph showed structural integrity in the tri-layer scaffold. In vivo data showed that the tri-layer scaffold accelerates wound healing in the mice model and angiogenesis in the chorioallantoic membrane model. Therefore, the designed scaffold inspired by the skin's structure can be used as a wound dressing to treat wounds.