Effect of treatment with phosphate, casein phosphopeptide and fluoride on the remineralization: in vitro study.

This study aimed to evaluate in vitro the effect protocols and anticaries agents containing casein amorphous calcium fluoride phosphopeptide-phosphate (CPP-ACPF, MI Paste Plus), sodium trimetaphosphate (TMP) and fluoride (F), in remineralization of caries lesions. Bovine enamel blocks with initial caries lesions were divided into groups (n = 12): 1) Toothpaste without F-TMP-MI Plus (Placebo); 2) Toothpaste 1100 ppm F (1100F), 3) 1100F + MI Paste Plus (1100F-MI Paste Plus), 4) Toothpaste with 1100F + Neutral gel with 4,500 ppm F + 5%TMP (1100F + Gel TMP) and 5) Toothpaste with 1100F + Neutral gel with 9,000 ppm F (1100F + Gel F). For the 4 and 5 groups the gel was applied only once for 1 minute, initially to the study. For the 3 group, after treatment with 1100F, MI Paste Plus was applied 2x/day for 3 minute. After pH cycling, the percentage of surface hardness recovery (%SHR); integrated loss of subsurface hardness (ΔKHN); profile and depth of the subsuperficial lesion (PLM); concentrations of F, calcium (Ca) and phosphorus (P) in enamel was determined. The data were analyzed by ANOVA (1-criterion) and Student-Newman-Keuls test (p < 0.001). Treatment with 1100F alone led to ~ 28% higher remineralization when compared to treatment with 1100F associated with MI Paste Plus (p < 0.001). The 1100F and 1100F + Gel F groups showed similar values for %SHR (p = 0.150). 1100F + Gel TMP treatment also remineralized the enamel surface by ~ 30% and 20% when compared to the 1100F + Gel F and 1100F groups (p < 0.001). The lower lesion depth (ΔKHN) was observed for the 1100F + Gel TMP group (p < 0.001), where it was 54% and 44% lower in comparison to the 1100F and 1100F + Gel F groups (p < 0.001). Polarized light microscopy photomicrographs showed subsurface lesions in all groups, but these lesions were present to a lower extent in the 1100F + Gel TMP group (p < 0.001). Treatment with 1100F + Gel TMP promoted an increase in the concentration of Ca in the enamel by ~ 57% and ~ 26% when compared to the 1100F and 1100F + MI Paste Plus groups (p < 0.001), respectively. There were no significant differences between the 1100F, 1100F + MI Paste Plus and 1100F + Gel F groups (p > 0.001). Similar values of P in the enamel were observed in the 1100F, 1100F + MI Paste Plus and 1100F + Gel F groups (p > 0.001), except for the 1100F + Gel TMP group, which presented a high concentration (p < 0.001). We conclude that the 1100F+TMP gel treatment/protocol led to a significant increased remineralization when compared to the other treatments/protocols and may be a promising strategy for patients with early caries lesions.

Evaluation of the cytotoxic activity of sorafenib-loaded camel milk casein nanoparticles against hepatocarcinoma cells.

Sorafenib, a multikinase inhibitor is used to treat hepatocellular and renal carcinoma. However, a low solubility impedes its bioavailability and thus, effectiveness. This study aims to enhance its effectiveness by using novel camel milk casein nanoparticles as a delivery system. This study evaluates the cytotoxicity of sorafenib encapsulated in camel milk casein nanoparticles against human hepatocarcinoma cells (HepG2 cells) in vitro. Optimal drug loaded nanoparticles were stable for 1 month, had encapsulation efficiency of 96%, exhibited a particle size of 230 nm, zeta potential of -14.4 and poly disparity index of 0.261. Treatment with it led to cell morphology and DNA fragmentation as a characteristic of apoptosis. Flow cytometry showed G1 phase arrest of cell cycle and 26% increased apoptotic cells population upon treatment as compared to control. Sorafenib-loaded casein nanoparticles showed 6-fold increased ROS production in HepG2 cells as compared to 4-fold increase shown by the free drug. Gene and protein expression studies done by qPCR and western blotting depicted upregulation of tumor suppressor gene p53, pro-apoptotic Bax, and caspase-3 along with downregulated anti-apoptotic Bcl-2 gene and protein expression which further emphasized death by apoptosis. It is concluded regarding the feasibility of these casein nanoparticles as a delivery system with enhanced therapeutic outcomes against hepatocellular carcinoma cells.

The effect of photobiomodulation therapy associated with casein phosphopeptide-amorphous calcium phosphate fluoride paste on the treatment of posthome whitening tooth sensitivity and color change: A randomized clinical trial.

OBJECTIVE: This study assessed whether combining photobiomodulation therapy (PBMT) with casein phosphopeptide-amorphous calcium phosphate fluoride (CPP-ACPF) paste can effectively reduce post-home whitening tooth sensitivity (TS) without compromising shade change. METHODS: Fifty participants were selected and assigned to one of four groups: (1) PLACEBO group-received a placebo paste and PBMT simulation; (2) PBMT group-received a placebo paste + PBMT; (3) CPP-ACPF group-received CPP-ACPF paste and PBMT simulation; (4) CPP-ACPF + PBMT group-received both CPP-ACPF paste and PBMT. The participants used whitening trays containing 22% carbamide peroxide for 2 h a day for 21 days. TS was measured daily using a visual analog scale, while shade change was assessed using a spectrophotometer: before bleaching treatment (T0), after the first (T1), second (T2), and third (T3) weeks of treatment, and 30 days (T4) after completing the whitening treatment. RESULTS: Intragroup analysis revealed that the PLACEBO group had the highest increase in sensitivity during the whitening treatment. The CPP-ACPF and PBMT groups showed no significant difference tooth whitening (TW) between weeks regarding aesthetic change. The CPP-ACPF and PBMT group exhibited a significant reduction in TS between the first and third and between the second and third weeks TW, but not between the first and second. Conversely, the PLACEBO group showed a higher sensitivity than the other groups (p < .05). The CPP-ACPF and PBMT groups did not differ from each other. Furthermore, the CPP-ACPF and PBMT group showed a greater decrease in sensitivity than the PLACEBO group at T1, T2, and T3 (p < .01), and was significantly differed from CPP-ACPF and PBMT groups only at T2 and T3. All groups confirmed TW effectiveness. Student's and paired t-test did not reveal any significant difference between groups (p > .05). CONCLUSION: Therefore, PBMT associated with CPP-ACPF paste can reduce TS without compromising the efficacy of TW.

Glutathione - IR 797 coupled Casein Nano-Trojan for augmenting the therapeutic efficacy of camptothecin in highly invasive triple negative breast cancer.

The rapid metastasis & heterogenic constitution of triple negative breast cancer (TNBC) limits drug entry to the tumor, reducing treatment effectiveness. To address this, we have synthesized Casein nanoparticles (Cn NPs) with attached glutathione (GSH), a natural ligand for cancer cell overexpressed γ-glutamyl transpeptidase (GGT). Cn NPs encapsulated with Camptothecin and NIR dye IR 797 (CCN NPs) for combinatorial therapy of TNBC. The GSH-CCN nanoparticles (CCNG NPs) act as a Nano-Trojan to deceive the cancer cells by delivering therapeutic payloads directly to specific target cells. In this study, Casein Nano-Trojan is equipped with GSH as a targeting ligand for GGT. The binding of CCNG NPs with cell surface receptors switched the anionic charge to catanionic, prompting the target cell to engulf the nanoparticles. The Casein Nano-Trojan releases its therapeutic payload inside the target cell, potentially inhibiting proliferation & inducing a high percentage of cell death (85 ± 7 %). Disintegration of mitochondrial membrane potential, inhibition of both migration & re-growth were observed. Immunofluorescence, acridine orange/ethidium bromide stain, and nuclear fragmentation assay further confirmed the substantial DNA damage induced by the high expression of γH2AX and p53. Significant therapeutic efficacy was observed in the 3D spheroids of 4T1 cells and in vivo breast cancer mice model (BALB/c). These findings demonstrate that CCNG NPs could be an effective treatment approach for highly metastatic triple negative breast cancer.

In-vitro effect of a single application of CPP-ACP pastes and different fluoridated solutions on the prevention of dental caries around orthodontic brackets.

OBJECTIVE: To assess the in-vitro effect of single applications of CPP-ACP pastes and different fluoridated solutions on the prevention of dental caries around orthodontic brackets. MATERIAL AND METHODS: Tooth/bracket sets (n=65) were immersed in artificial saliva (1h at 37ºC) and randomly subjected to single applications (100µL; 1min) of casein phosphopeptide-amorphous calcium phosphate (CPP-ACP emulsion), CPP-ACP with fluoride (CPP-ACPF emulsion), solutions of titanium tetrafluoride (TiF4) or sodium fluoride (NaF), or no treatment (CG). Multispecies biofilm (5 x 105 CFU/mL) was formed in the presence of 2% sucrose. After 24 h, the pH and the concentration of total soluble fluoride (TSF) were analyzed by culture medium. The presence of active white spot lesions (WSL) evaluated by macroscopic examination and the percent surface mineral loss (%SML) were analyzed. Also, the topography of enamel was detected by analysis of scanning electron microscopy (SEM). The data was assessed by chi-square, Kruskal-Wallis, and Mann-Whitney tests (p < 0.05). RESULTS: Fluoride-containing compounds led to a smaller pH reduction than did CPP-ACP and CG (p<0.05). There was difference in TSF between the groups (p<0.05), denoted as TiF4> NaF > CPP-ACPF > CPP-ACP > CG. Regarding the presence of WSL and %SML, the NaF group obtained lower values (p<0.05), while TiF4 and CPP-ACPF were similar (p>0.05). SEM demonstrated that fluoride-free groups had a larger surface dissolution. CONCLUSION: Fluoridated groups including solutions and CPP-ACPF were more effective than CPP-ACP in reducing enamel demineralization around orthodontic brackets after a single application.

An in vitro evaluation of the effects of fluoride, CPP-ACP, or resin infiltration on discoloration caused by pediatric supplements.

OBJECTIVES: White spot lesions are more susceptible to staining agents due to their porous structure. This study examines the impact of white spot lesion treatments on discoloration caused by pediatric supplements. METHOD AND MATERIALS: Three treatments (fluoride, casein phosphopeptide-amorphous calcium phosphate [CPP-ACP], resin infiltration), a control, and their respective syrup-based subgroups (iron and black elderberry syrups) were established, each with eight teeth. Artificial white spot lesions were induced, and weekly applications of fluoride varnish, daily applications of CPP-ACP paste, or a single resin infiltration procedure were performed on the white spot lesions within the treatment groups over 4 weeks. Simultaneously, samples were exposed daily to iron or black elderberry syrups. Spectrophotometer measurements were taken at baseline, after demineralization (T0), and after 1 (T1), 2 (T2), and 4 weeks (T4). ΔE00 values were calculated. Statistical analysis was conducted using a three-way mixed-design ANOVA, with the significance level set at P = .05. RESULTS: At T4, ΔE00 values from all groups exceeded the clinical acceptability limit of 1.8. At T2 and T4, the ΔE00 values obtained from the black elderberry syrup subgroups were significantly higher (P < .001). At T4, the highest ΔE00 values were seen in the CPP-ACP groups (P < .001). The lowest ΔE00 values at T2 and T4 were observed in the resin infiltration groups (P < .05). CONCLUSIONS: Supplements containing ferrous sulfate and black elderberry extract caused color changes in white spot lesions that exceeded the clinical acceptability limit. Resin infiltration of white spot lesions provides advantages over remineralization treatments, particularly in minimizing discoloration induced by pediatric supplements.

Combined Casein Phosphopeptide-Amorphous Calcium Phosphate and Fluoride Is Not Superior to Fluoride Alone in Early Carious Lesions: A Meta-Analysis.

There is a growing need for effective methods in the management of early stage carious lesions. Therefore, the aim of this study was to evaluate the effect of combined casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) and fluoride on white spot lesions (WSLs) compared to fluoride-only interventions. This meta-analysis was performed according to PRISMA guidelines and registered in PROSPERO (CRD42021286245). The Medline, Embase, and Cochrane Central databases were searched until October 17, 2022. Eligible studies were randomized controlled trials. Outcome variables included laser fluorescence (LF), quantitative light-induced fluorescence (QLF), and lesion area scores. The random-effects model was used for analysis, and results were given as standardized mean difference (SMD) and mean difference (MD) with a 95% confidence interval. Risk of bias was assessed using the RoB 2 tool, and the level of evidence with GRADE. Our systematic search yielded 973 records after duplicate removal, 21 studies were included for qualitative synthesis, and 15 studies were eligible for quantitative analysis. No significant difference was found between CPP-ACP and fluoride versus fluoride alone in LF at 1, 3, and 6 months of use: SMD -0.30 (-0.64; 0.04); SMD -0.47 (-1.02; 0.07); SMD -0.49 (-1.13; 0.15), respectively. For QLF, the analysis did not demonstrate significant differences between these two kinds of treatment at 1 and 6 months of use: MD 0.21 (-0.30;0.71); MD 0.60 (-1.70;2.90), but at 3 months, higher QLF values were found in the fluoride-only group compared to the CPP-ACP and fluoride combination was shown regarding the WSLs: MD 0.58 (0.25;0.91). On the contrary, data showed a small but statistically significant decrease in the lesion area in favor of the CPP-ACP plus fluoride versus fluoride alone at 6 months MD -0.38 (-0.72; -0.04). None of these observed changes indicated substantial clinical relevance. The combination of CPP-ACP and fluoride did not overcome the effect of fluoride given alone. Our data suggest that fluoride itself is effective in improving WSLs. However, the certainty of evidence was very low. These results indicate that further studies and future development of more effective products than CPP-ACP are needed in addition to fluoride to achieve robust amelioration of WSLs.

Estimation of the Efficacy of Remineralizing Agents on the Microhardness of Deciduous Teeth Demineralized Using Pediatric Formulations.

AIM: This study aimed to evaluate the demineralizing effect of commonly used pediatric syrup formulations on primary teeth and the efficacy of two readily available remineralizing agents in treating this effect. MATERIALS AND METHODS: Ninety primary teeth were used for sample preparation and divided into three groups: antibiotic syrup (group A), cough syrup (group B), and control (group C) groups. These groups were further categorized into intragroups according to the treatment with remineralizing agents: groups A1, B1, and C1 received GC Tooth Mousse (casein phosphopeptide-amorphous calcium phosphate, CPP-ACP paste) and groups A2, B2, and C2 received Clinpro Tooth Crème. The samples were subjected to a series of demineralization cycles for 14 days, and remineralization cycles until 30 days were performed using two remineralizing agents, that is, GC Tooth Mousse (CPP-ACP paste) and Clinpro Tooth Crème and were evaluated using Vicker's microhardness test. RESULTS: Antibiotic syrup (group A) and cough syrup (group B) showed a significant decrease in surface microhardness compared with control (group C). All intragroups showed an increase in surface microhardness after treatment with remineralizing agents, which was significantly higher in intragroups A1, B1, and C1 treated with GC Tooth Mousse (CPP-ACP paste). CONCLUSIONS: Oral liquid medications showed definite demineralization potential. CPP-ACP paste was found to be better than Clinpro Tooth Crème for demineralized teeth. CLINICAL SIGNIFICANCE: The use of over-the-counter drugs has increased among the average Indian population, especially for the treatment of fever, cold, and cough. Unwise use of medications by the present population without proper medical guidance will lead to irreparable changes in future generations.

Evaluating the Potential of Casein Glycomacropeptide in Adult Irritable Bowel Syndrome Management: A Pilot Study.

Irritable bowel syndrome (IBS) is a common gastrointestinal disorder that affects 10-15% of the global population and presents symptoms such as abdominal discomfort, bloating and altered bowel habits. IBS is believed to be influenced by gut microbiota alterations and low-grade inflammation. Bovine kappa-casein glycomacropeptide (GMP), a bioactive dairy-derived peptide, possesses anti-adhesive, prebiotic and immunomodulatory properties that could potentially benefit IBS patients. This pilot study investigated the effects of daily supplementation with 30 g of GMP for three weeks on gut health in five people with IBS. We assessed alterations in gut microbiota composition, fecal and blood inflammatory makers, and gut-related symptoms before, during and after the GMP feeding period. The results revealed no changes in fecal microbiota, subtle effects on systemic and intestinal immune makers, and no changes in gut-related symptoms during and after the GMP supplementation. Further research is needed to assess the potential benefits of GMP in IBS patients, including the examination of dosage and form of GMP supplementation.

Dentine tubule occlusion effect of hydrolyzed casein in a bioactive glass-based dental desensitizing gel.

OBJECTIVES: The effectiveness of three different groups of polyethylene glycol (PEG)-based gels containing powders on dentin hypersensitivity (DH) treatment were assessed and compared with Actimins® as commercial reference group. METHODS: Hydroxyapatite nanorods (nHA) and sol-gel-derived 45S5 bioglass (SGD 45S5) powders were synthesized through hydrothermal and sol-gel methods, respectively. First, 25 demineralized dentin disks were divided into five groups. Then, the prepared gels based on 45S5 bioglass with and without hydrolyzed casein (HC) as experimental, nHA gel and Actimins® as positive and commercial reference groups were applied twice a day on disks by a micro applicator. To mimic the oral environment, treated disks were immersed in artificial saliva in a water bath at 37 °C for 7 days. However, in the negative control group, no agent was applied on the samples. FE-SEM, EDS, AFM, and XRD were performed to assess tubule occlusion. One-way ANOVA test was used for statistical analysis and p*<0.05 was set as the significance level. RESULTS: The nHA with an average aspect ratio of 2.77 and the SGD 45S5 powders with a polygonal morphology and the average size of 48.64±11.38 µm were synthesized. After treatment, tubule occlusion in HC-SGD 45S5 and nHA gels were shown to be higher than other groups. The root mean square roughness (Rrms) of the above-mentioned gels showed to be 121.54±9.25 nm, and 312.6 ± 9 nm, respectively. CONCLUSION: The nHA containing group exhibited the highest tubule occlusion efficiency (i.e., tubule diameter of 0.92±0.32 µm) with a superior mineral precipitation. HC as a novel material demonstrates to be potentially beneficial in DH treatment. CLINICAL SIGNIFICANCE: DH as a common issue may be reduced or eliminated by occlusion of patent dentinal tubules. There are various types of desensitizing agents capable of controlling the DH by the occlusion of patent dentinal tubules. The desensitizing gels developed in this study showed to be promising for clinical and home-use applications.

Effect of three different remineralising agents on prevention against acidic erosion of primary teeth: an in vitro study.

PURPOSE: This study aimed to evaluate and compare the protective effect of fluoride varnish (Enamelast™, Ultradent Inc., Cologne, Germany), casein phosphopeptide-amorphous calcium phosphate fluoride/CPP-ACPF (MI Paste Plus, GC Corp., Tokyo, Japan) and self-assembling P11-4 peptide (Curodont™ Protect, Credentis AG, Windisch, Switzerland), against acidic erosion of primary teeth. METHODS: Forty primary anterior teeth were randomly assigned to four groups (n = 10): group 1: control, group 2: fluoride varnish, group 3: CPP-ACPF and group 4: self-assembling P11-4 peptide. After applying remineralising agents, except for the control group, all specimens underwent an erosive challenge of carbonated soft drink and artificial saliva for 15 cycles of 6 s each at 6-h intervals for a day. Groups were compared in terms of surface microhardness, roughness readings, and surface scanning with an extra-oral scanner (D800; 3Shape A/S) before and after the erosive process. RESULTS: All experimental groups showed superior results than the control group regarding microhardness, surface roughness, and substance loss. The fluoride varnish group showed significantly favourable results in microhardness change. There was no significant difference between the experimental groups regarding surface roughness and enamel loss measurements. CONCLUSION: 5% NaF fluoride varnish, CPP-ACPF and self-assembling P11-4 peptide protect the enamel of primary teeth against erosion compared to artificial saliva alone.

Shear bond strength of metallic brackets bonded to enamel pretreated with CPP-ACP: a systematic review and meta-analysis of in vitro studies.

BACKGROUND: Development of white spot lesions (WSLs) is common among orthodontic patients. Several measures have been introduced to prevent and remineralize the lesions. Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) is used for both prevention and remineralization. The effect of its application before bonding is controversial. This systematic review was conducted to investigate the most up to date available literature regarding the effect of CPP-ACP enamel pre-treatment on shear bond strength (SBS) of metallic orthodontic brackets. METHODS: A search was conducted in electronic databases (MEDLINE (via PubMed), Scopus, Cochrane Library, Web of Science and Google scholar (grey literature)) up to March 29th, 2023. The inclusion criteria included in vitro studies comparing the SBS of metal orthodontic brackets following pre-treatment of enamel using CPP-ACP versus control. The exclusion criteria included study types other than in vitro studies, studies conducted on non-human enamel, or studies using CPP-ACP in combination with another intervention. The included studies were analysed by two reviewers, independently. The risk of bias assessment was done using a modified risk of bias tool. A Meta-analysis was performed. I2 values and Q-test were used for assessment of heterogeneity. Results were displayed in forest plots with a random-effects model. Standardized mean difference, standard error (SE) and 95% confidence intervals were calculated for all studies. RESULTS: The search resulted in 76 articles. After duplicate removal and assessment for eligibility, 15 studies were included in the review. High statistical heterogeneity was found among the included studies using I2 values and Q-Test (I2 = 95.147%; Q = 288.456; df = 14; P < 0.001). The overall effect of CPP-ACP pre-treatment on the SBS of metal orthodontic brackets was not significant (Mean difference = 1.163 MPa, SE = 0.757, 95% CI = -0.321, 2.648, p value = 0.125). The use of CPP-ACP for prevention of WSLs did not significantly affect the SBS of brackets (Standardized mean difference = 1.009, SE = 0.884, 95% CI = -0.723, 2.740, p value = 0.254). No significant change was found when CPP-ACP was used for remineralization of WSLs (Standardized mean difference = 1.501, SE = 1.087, 95% CI = -0.630, 3.632, p value = 0.167). CONCLUSIONS: Within the limitations of the study, the evidence suggests that the use of CPP-ACP for either prevention or remineralization of WSLs before bonding does not affect the SBS of metal orthodontic brackets.

Comparison of therapies of white spot lesions: a systematic review and network meta-analysis.

OBJECTIVE: White spot lesions (WSLs), the earliest evidence of enamel demineralization, are considered amenable to intervention to achieve a remineralized or arrested state of caries. The management of WSLs is quite challenging, and there is no definitive cure as yet. We performed a network meta-analysis to assess the efficacy of seven therapies for WSLs and gave a hierarchy of them. MATERIALS AND METHODS: We systematically searched the PubMed, EMBASE, Cochrane, and Web of Science databases (last search: July 2022) to identify all relevant studies. We limited our search to studies published in English. Randomized controlled designed in vitro/clinical trials related to the efficacy of the seven therapies for WSLs were included. Data extraction was performed independently by two reviewers. The risk of bias (ROB) 2.0 tool from Cochrane and a previous in vitro methodological tool will be used for the quality assessment. Variations in quantitative light-induced fluorescence (QLF), laser fluorescence (LF), and lesions area were the primary outcome measures. Standard mean difference (SMD) was used as the effect size for the Network meta-analysis (NMA). Consistency and inconsistency tests were conducted. The hierarchy of 7 treatment effects was evaluated using surface probabilities under cumulative ranking (SUCRA). Publication bias was evaluated using a bias plot. RESULTS: Forty-two articles were included in the systematic review. Thirty-one of them, with a total of 1906 participants, were included in the network meta-analysis. The studies owned a low and moderate risk of bias. This analysis does not suffer from significant inconsistency. The difference between 4 groups 'self-assembled peptide (SAP) P11-4', 'P11-4 + Fluoride Varnish (FV)', 'Resin Infiltration (RI)', 'casein phosphor peptides-amorphous calcium fluoride phosphate (CPP-ACFP)' and the 'Control' group was found to be statistically significant. Compared to the 'FV' and 'casein phosphor peptides-amorphous calcium phosphate (CPP-ACP)' groups, the 'P11-4 + FV" group and 'RI" group made a significant difference. The hierarchy was evident in the SUCRA values of 7 therapies. P11-4 + FV and RI were considered effective therapies compared to the control group or the FV group (gold standard group). CONCLUSIONS: The available evidence suggests that resin infiltration and P11-4 in combination with fluoride varnish had advantages over gold standard (FV). The effect of tricalcium phosphate-based drugs and fluoride is not very noticeable. Overall, drugs based on P11-4 and resin infiltration will be better therapies. Using more than two drugs in combination also would increase efficacy.

An active new formulation of iron carried by aspartyl casein for iron-deficiency anemia: results of the ACCESS trial.

Oral iron supplementation is the cornerstone for the management of iron-deficiency anemia. A new oral formulation of iron conjugated with N-aspartyl-casein (Fe-ASP) (Omalin®, Uni-Pharma) is studied in the ACCESS double-blind, double-dummy randomized clinical trial; 60 patients were randomized to 12-week oral treatment twice every day either with oral ferrous sulfate (FeSO4) delivering 47 mg elementary iron or oral Fe-ASP delivering 40 mg elementary iron. Participants had hemoglobin less than 10 g/dl, decreased red blood cell (RBC) count, and ferritin lower than 30 ng/ml; patients with a medical history of malignancy were excluded. The primary endpoint was the increase of Hb in the first 4 weeks of treatment, and the study was powered for non-inferiority. A new score of global improvement was introduced where all participants were given one point for any at least 10% increase of Hb, RBC, and reticulocytes. At week 4, the mean (SE) change of Hb was 0.76 g/dl in the FeSO4 group and 0.83 g/dl in the Fe-ASP group (p: 0.876). The odds for worse allocation of the global score were 0.35 in the Fe-ASP group compared to the FeSO4 group. Patients in the Fe-ASP group experienced a significant decrease in the number of IDA-related physical signs by week 4. No differences were found between the two groups in any of the patient-reported outcomes of fatigue and of gastrointestinal adverse events either at week 4 or at week 12. ACCESS is the most recent clinical trial showing the non-inferiority of Fe-ASP to FeSO4 for the primary endpoint of the Hb change.

Diffusion-mediated carving of interior topologies of all-natural protein nanoparticles to tailor sustained drug release for effective breast cancer therapy.

Proteins are promising base materials for developing drug carriers with efficient blood circulation due to low possibilities of clearance by macrophages. However, such natural biopolymers have highly sophisticated molecular structures, preventing them from being assembled into nano-platforms with manipulable payload release profiles. Here, we report the self-assembly of two natural proteins (milk casein and rice protein) into protein nanoparticles (NPs, ∼150 nm) with tailorable release profiles. Diffusion of plant-derived paclitaxel (PTX)-containing eugenol into the hydrophobic cores of the NPs and subsequent dialysis to remove eugenol from the cores lead to the carving of the NP interiors. With the increase in the mass ratios of casein and rice protein, this process generates all-natural NPs with PTX loaded in their full cavities, semi-full cavities, or solid cores. These NPs can be efficiently uptaken by breast cancer cells and could kill the cancer cells efficiently. PTX in these NPs demonstrates increasingly sustained in vivo release profiles from full cavities, semi-full cavities, to solid cores, gradually extending its pharmacokinetic profiles in blood plasma to favor drug accumulation in breast tumor models. Consequently, the NPs with solid cores completely inhibit tumor growth in vivo, more effectively than those with full and semi-full cavities. Our work opens up a new avenue to the use of diffusion-mediated nanoscale carving in producing biomaterials with controllable interior topologies relevant to drug release profiles.

Effects of milk protein concentrate supplementation on metabolic parameters, adipocytokines and body composition in obese women under weight-loss diet: study protocol for a randomised controlled trial.

INTRODUCTION: Obesity impairs metabolic function and increases the risk of cardiovascular disease and type 2 diabetes mellitus. Evidence suggests that high-protein diets help to increase weight loss and protect against weight gain. Milk protein concentrate (MPC) is a dairy product with a high protein content with a ratio of casein and whey protein similar to skim milk. This trial aims to evaluate the effect of MPC supplementation in obese women under a weight-loss diet. METHODS AND ANALYSIS: We will conduct a 2-month open-label, parallel-group, randomised controlled trial to determine the effect of MPC supplementation on levels of glycaemic and lipid profile, leptin, adiponectin, appetite, waist circumference, body mass index and body composition in 44 premenopausal obese women on a weight-loss diet. ETHICS AND DISSEMINATION: This protocol, approved by the Medical Ethics Committee of Ahvaz University of Medical Sciences, is in accordance with the Declaration of Helsinki (approval number: IR.AJUMS.REC.1399.795). The trial results will be published in peer-reviewed journals. TRIAL REGISTRATION NUMBER: Iranian Registry of Clinical Trials (IRCT20201223049804N1).

Molecular Characterization of Probiotics and Their Influence on Children with Autism Spectrum Disorder.

Children with autism spectrum disorder (ASD) are usually unable to express abdominal discomfort properly, and thus gastrointestinal symptoms (GIS) are sometimes shadowed by aggression, which is sometimes misunderstood as a behavioral characteristic of ASD. Several studies have reported interesting correlations between the severity of behavioral and gastrointestinal symptoms in ASD children. The present study aimed to investigate the potential effects of probiotics as an adjuvant therapy to modulate the clinical status of ASD children. This study included 40 children with ASD aged 2-5 years. The feeding product was prepared from whey powder (without casein) and some minced cooked yellow vegetables in adequate ratios fortified with the studied probiotic strains (Bifidobacterium spp. and Lactobacillus spp.). Bifidobacterium strains were assessed from stool samples of children with ASD. Bifidobacterium strains were analyzed in the stools of ASD children. Recruited ASD patients received 10 g of the nutritional supplement once a day for 3 months. Childhood Autism Rating Scale (CARS) and Autism Diagnostic Interview-Revised (ADIR) were reevaluated clinically. Questionnaire on Pediatric Gastrointestinal Symptoms-Rome III Version was used for all children with ASD before and after. There is a significant increase in the colony counts of both Bifidobacterium spp. and Lactobacillus spp., which present in the stool of ASD children after probiotic supplementation for 3 months. It was highly significant in the case of Bifidobacterium spp. (p value 0.000) and a significant increase in Lactobacillus spp. (p value 0.015). The present study showed reduced anxiety and observation of deep sleep for children with ASD (80%) after taking the supplementation. This indicates that probiotics may have a potential effect in reducing symptoms and severity of ASD and in correcting dysbiosis.

RNA sequencing and bioinformatics analysis revealed PACSIN3 as a potential novel biomarker for platinum resistance in epithelial ovarian cancer.

BACKGROUND: Failure to respond to treatment in epithelial ovarian cancer can often be attributed to platinum-based chemotherapy resistance. However, the possible mechanisms or candidate biomarkers associated with platinum resistance are yet to be elucidated, even though many researchers have performed related studies. METHODS: We performed RNA sequencing of clinical specimens obtained from patients with platinum-sensitive or resistant epithelial ovarian cancer (EOC). Furthermore, various bioinformatics approaches, including spatial analysis of functional enrichment, were used to identify key regulators and associated underlying mechanisms of platinum resistance in EOC. RESULTS: Through RNA-sequencing, we identified 263 differentially expressed genes (98 upregulated and 165 downregulated) and subjected them to Gene Oncology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses, which were characterized to the traditional platinum-resistant characteristics. Subsequently, the gene interaction network and module analysis by spatial analysis of functional enrichment software demonstrated protein kinase C and casein kinase substrate in neurons 3 (PACSIN3) as the only upregulated hub gene, and neurotensin (NTS) and KIAA0319 as downregulated hub genes in platinum-resistant EOC. We selected PACSIN3 for further analysis because it has not been studied in relation to response to platinum-based chemotherapy. PACSIN3 was significantly upregulated in ovarian cancer cells compared to immortalized human ovarian surface epithelial cells. In addition, cisplatin-induced apoptosis was measured in PACSIN3 knockout OVCA433 and BRCA-mutated EOC cell line, SNU251, by a fluorescence-activated cell sorting-based Annexin-V/propium iodide double staining assay, which revealed a significant increase in apoptosis. CONCLUSIONS: Taken together, the present study presents PACSIN3 as a promising predictive biomarker associated with platinum resistance, especially in BRCA-mutated epithelial ovarian cancers.

Comparative evaluation of remineralisation potential of bioactive glass, casein phosphopeptide-amorphous calcium phosphate and novel strontium-doped nanohydroxyapatite paste: An In-Vitro study.

Background: Many studies explained the importance of remineralisation of early carious lesions with various remineralising agents. In the present study, we incorporated the remineralising agents in a dentifrice, applied that in artificial enamel caries and evaluated their remineralising potential and compared the efficacy among the three. Aim: To evaluate and compare the remineralisation potential of a dentifrice containing bioactive glass, casein phosphopeptide-amorphous calcium phosphate and novel laboratory synthesised strontiumdoped nanohydroxyapatite paste in artificial enamel caries. Methods and Materials: 120 enamel specimens were divided into 4 groups of 30 specimens each, based on the type of dentifrice applied: GI - conventional toothpaste (control group), GII - calcium sodium phosphosilicate (Novamin), GIII - casein phosphopeptide-amorphous calcium phosphate (GC tooth mousse) and GIV- Novel strontiumdoped nanohydroxyapatite paste (SrnHAp paste). Specimens in all the groups were subjected to demineralisation, and calcium/phosphorous ratio was analysed followed by remineralisation and the mean calcium-phosphorus ratio was assessed using a scanning electron microscope and energy dispersing X-ray analysis. Statistical Analysis: Data were analysed using IBM SPSS Statistics for Windows Software, version 22 (IBM Corp., Armonk, NY, USA). Descriptive statistics were used to calculate the mean and standard deviation. Kruskal-Wallis, ANOVA and Mann-Whitney tests were used. The level of significance was set at 5%. Results and Conclusion: All except the control group showed a net increase in calcium and phosphorous values after application of the respective remineralising agents in respective groups. Inter-group comparison revealed that Group IV - SrnHAp paste yields higher net calcium and phosphorous values than other groups. Hence, novel SrnHAp can be considered as the material of choice in remineralising early enamel carious lesions.

Pharmaceutical Potential of Casein-Derived Tripeptide Met-Lys-Pro: Improvement in Cognitive Impairments and Suppression of Inflammation in APP/PS1 Mice.

BACKGROUND: Tripeptide Met-Lys-Pro (MKP), a component of casein hydrolysates, has effective angiotensin-converting enzyme (ACE) inhibitory activity. Brain angiotensin II enzyme activates the NADPH oxidase complex via angiotensin II receptor type 1 (AT1) and enhances oxidative stress injury. ACE inhibitors improved cognitive function in Alzheimer's disease (AD) mouse models and previous clinical trials. Thus, although undetermined, MKP may be effective against pathological amyloid-ß (Aß) accumulation-induced cognitive impairment. OBJECTIVE: The current study aimed to investigate the potential of MKP as a pharmaceutical against AD by examining MKP's effect on cognitive function and molecular changes in the brain using double transgenic (APP/PS1) mice. METHODS: Experimental procedures were conducted in APP/PS1 mice (n = 38) with a C57BL/6 background. A novel object recognition test was used to evaluate recognition memory. ELISA was used to measure insoluble Aß40, Aß42, and TNF-α levels in brain tissue. Immunohistochemical analysis allowed the assessment of glial cell activation in MKP-treated APP/PS1 mice. RESULTS: The novel object recognition test revealed that MKP-treated APP/PS1 mice showed significant improvement in recognition memory. ELISA of brain tissue showed that MKP significantly reduced insoluble Aß40, Aß42, and TNF-α levels. Immunohistochemical analysis indicated the suppression of the marker for microglia and reactive astrocytes in MKP-treated APP/PS1 mice. CONCLUSION: Based on these results, we consider that MKP could ameliorate pathological Aß accumulation-induced cognitive impairment in APP/PS1 mice. Furthermore, our findings suggest that MKP potentially contributes to preventing cognitive decline in AD.