Cytotoxic Activity of Non-specific Lipid Transfer Protein (nsLTP1) from Ajwain \u3cem\u3e(Trachyspermum ammi)\u3c/em\u3e Seeds

Background Trachyspermum ammi, commonly known as Ajwain, is a member of the Apiaceae family. It is a therapeutic herbal spice with diverse pharmacological properties, used in traditional medicine for various ailments. However, all previous studies were conducted using small molecule extracts, leaving the protein’s bioactivity undiscovered. Aim The current study aimed to demonstrate the cytotoxic activity of Ajwain non-specific lipid transfer protein (nsLTP1) in normal breast (MCF10A), breast cancer (MCF-7), and pancreatic cancer (AsPC-1) cell lines. Also, to evaluate its structural stability in human serum as well as at high temperature conditions. Methods The cytotoxic activity of Ajwain nsLTP1 was evaluated in MCF-7 and AsPC-1 cell lines using MTT assay. Annexin V-FITC and PI staining were used to detect the early apoptotic and late apoptotic cells. The role of nsLTP1 in inducing apoptosis was further studied by quantifying Bcl-2, Bax, Caspase-3, Survivin, EGFR, and VEGF genes expression using RT-PCR. CD spectroscopy analyzed the nsLTP1 conformational changes after thermal treatment for structure stability determination. The RP-HPLC was used to analyze the nsLTP1 degradation rate in human serum at different time intervals incubated at 37 °C. Results Ajwain nsLTP1 showed a potent cytotoxic effect in MCF-7 and AsPC-1. The IC50 value obtained in MCF-7 was 8.21 μM, while for AsPC-1 4.17 μM. The effect of nsLTP1 on stimulating apoptosis revealed that the proportions of apoptotic cells in both cell lines were relatively increased depending on the concentration. The apoptotic cells percentage at 20 μM was in MCF-7 71% (***P \u3c 0.001) and AsPC-1 88% (***P \u3c 0.001). These results indicate that nsLTP1 might efficaciously induce apoptosis in multiple types of cancerous cells. Genes expression in MCF-7 and AsPC-1 showed significant upregulation in Bax and Caspase-3 and downregulation in Bcl-2, Survivin, EGFR, and VEGF protein. The CD analysis of nsLTP1 showed a significant thermostable property. In serum, nsLTP1 showed a slow degradation rate, indicating high stability with a half-life of ~ 8.4 h. Conclusion Our results revealed the potential anticancer activity of Ajwain nsLTP1 and its mechanism in inducing apoptosis. It further exhibited thermostable properties at high temperatures and in human serum, which suggested this protein as a promising anticancer agent

The Structural Characterization and Bioactivity Assessment of Nonspecific Lipid Transfer Protein 1 (nsLTP1) from Caraway (\u3cem\u3eCarum carvi\u3c/em\u3e) Seeds

Background Carum carvi (caraway) of the Apiaceae family has been used in many cultures as a cooking spice and part of the folk medicine. Previous reports primarily focus on the medicinal properties of caraway seed essential oil and the whole seeds extract. However, no effort has been made to study caraway proteins and their potential pharmacological properties, including nonspecific lipid transfer protein (nsLTP), necessitating further research. The current study aimed to characterize nonspecific lipid transfer protein 1 (nsLTP1) from caraway seed, determine its three-dimensional structure, and analyze protein–ligand complex interactions through docking studies. We also evaluated nsLTP1 in vitro cytotoxic effect and antioxidant capacity. Additionally, nsLTP1 thermal- and pH- stability were investigated. Methods Caraway nsLTP1 was purified using two-dimensional chromatography. The complete amino acid sequence of nsLTP1 was achieved by intact protein sequence for the first 20 residues and the overlapping digested peptides. The three-dimensional structure was predicted using MODELLER. Autodock Vina software was employed for docking fatty acids against caraway nsLTP1. Assessment of nsLTP1 cytotoxic activity was achieved by MTS assay, and the Trolox equivalent antioxidant capacity (TAC) was determined. Thermal and pH stability of the nsLTP1 was examined by circular dichroism (CD) spectroscopy. Results Caraway nsLTP1 is composed of 91 residues and weighs 9652 Da. The three-dimensional structure of caraway nsLTP1 sequence was constructed based on searching known structures in the PDB. We chose nsLTP of Solanum melongena (PDB ID: 5TVI) as the modeling template with the highest identity among all other homologous proteins. Docking linolenic acid with caraway protein showed a maximum binding score of -3.6 kcal/mol. A preliminary screening of caraway nsLTP1 suppressed the proliferation of human breast cancer cell lines MDA-MB-231 and MCF-7 in a dose‑dependent manner with an IC50 value of 52.93 and 44.76 μM, respectively. Also, nsLTP1 (41.4 μM) showed TAC up to 750.4 μM Trolox equivalent. Assessment of nsLTP1 demonstrated high thermal/pH stability. Conclusion To the best of our knowledge, this is the first study carried out on nsLTP1 from caraway seeds. We hereby report the sequence of nsLTP1 from caraway seeds and its possible interaction with respective fatty acids using in silico approach. Our data indicated that the protein had anticancer and antioxidant activities and was thermally stable

Repurposing tofacitinib as an anti-myeloma therapeutic to reverse growth-promoting effects of the bone marrow microenvironment.

The myeloma bone marrow microenvironment promotes proliferation of malignant plasma cells and resistance to therapy. Activation of JAK/STAT signaling is thought to be a central component of these microenvironment-induced phenotypes. In a prior drug repurposing screen, we identified tofacitinib, a pan-JAK inhibitor Food and Drug Administration (FDA) approved for rheumatoid arthritis, as an agent that may reverse the tumor-stimulating effects of bone marrow mesenchymal stromal cells. Herein, we validated in vitro, in stromal-responsive human myeloma cell lines, and in vivo, in orthotopic disseminated xenograft models of myeloma, that tofacitinib showed efficacy in myeloma models. Furthermore, tofacitinib strongly synergized with venetoclax in coculture with bone marrow stromal cells but not in monoculture. Surprisingly, we found that ruxolitinib, an FDA approved agent targeting JAK1 and JAK2, did not lead to the same anti-myeloma effects. Combination with a novel irreversible JAK3-selective inhibitor also did not enhance ruxolitinib effects. Transcriptome analysis and unbiased phosphoproteomics revealed that bone marrow stromal cells stimulate a JAK/STAT-mediated proliferative program in myeloma cells, and tofacitinib reversed the large majority of these pro-growth signals. Taken together, our results suggest that tofacitinib reverses the growth-promoting effects of the tumor microenvironment. As tofacitinib is already FDA approved, these results can be rapidly translated into potential clinical benefits for myeloma patients

Influence of additives (inorganic/organic) on the clouding behavior of amphiphilic drug solutions: Some thermodynamic studies

AbstractHerein we provide a detailed result about the effect of various additives, viz. inorganic salts, quaternary ammonium bromides (QABs) and amino acids on clouding behavior of amphiphilic drug amitriptyline hydrochloride (AMT). The continuous increase in the cloud point (CP) of drug by increase in inorganic salt concentration and the magnitude of increases rely upon the position of the salts in Hofmeister series and hydrated radii. The QABs also influence continuous increase in the CP, which is illustrated in terms of the alkyl chain length of peculiar QAB. The effect of amino acids on CP of the drug solution is dependent upon the characteristics (acidic, basic, polar or nonpolar) of particular amino acids. The overall behavior of additives has been analyzed and discussed on the basis of electrostatic repulsion or interaction, micellar growth, and mixed micelle formation between the ingredients. In addition to this, thermodynamic parameters are also evaluated

Impact of Pediatric Obesity on Diurnal Blood Pressure Assessment and Cardiovascular Risk Markers

Background: The prevalence of hypertension is increasing particularly among obese children and adolescents. Obese children and adolescents with hypertension are likely to remain hypertensive as they reach adulthood and hypertension is linked to an increased risk for cardiovascular disease. Twenty-four-hour ambulatory blood pressure monitoring (ABPM) has become one of the most important tools in diagnosing hypertension in children and adolescents and circadian patterns of blood pressure may be important disease-risk predictors. Methods: A retrospective chart review was conducted in patients aged 6–21 years who underwent 24-h ABPM at Kentucky Children\u27s Hospital (KCH) from August 2012 through June 2017. Exclusion criteria included conditions that could affect blood pressure including chronic kidney disease and other renal abnormalities, congenital heart disease, cancer, and thyroid disease. Subjects were categorized by body mass index into normal (below 85th percentile), overweight (85th−95th percentile), stage I obesity (95th−119th percentile), stage II obesity (120th−139th) and stage III obesity (\u3e 140th). Non-dipping was defined as a nocturnal BP reduction of \u3c 10%. Results: Two hundred and sixty-three patients (156 male patients) were included in the analysis, of whom 70 were normal weight, 33 overweight, 55 stage I obesity, 53 stage II, and 52 stage III obesity. Although there was no significant difference between normal weight and obese groups for prevalence of hypertension, there was a greater prevalence of SBP non-dipping in obese patients as BMI increased (p = 0.008). Furthermore, non-dippers had a significantly elevated LVMI as well as abnormal lab values for uric acid, blood lipid panel, creatinine, and TSH (p \u3c 0.05). Conclusions: These findings demonstrate that obese children and adolescents constitute a large proportion of hypertensive children and adolescents and the severity of pediatric obesity is associated with nocturnal BP non-dipping. Additionally, obesity in children is linked to several cardiovascular risk factors including left ventricular hypertrophy, dyslipidemia, and elevated uric acid levels. Further studies utilizing ABPM measures on risk stratification in this very high-risk population are warranted

Clinical performance and Willingness To Pay for soft toric contact lenses in low and moderate astigmats

Purpose: To determine clinical performance and the ‘Willingness To Pay’ for toric vs. spherical soft contact lenses in an astigmatic population. Methods: In the clinical study, subjects with binocular low to moderate astigmatism (-0.75DC to − 1.50DC) wore pairs of soft toric (Biofinity toric) and spherical (Biofinity) contact lenses in random sequence. Visual acuity (high and low contrast, monocular and binocular), subjective comfort and subjective vision were recorded. In the economics study, first subjects who had participated in the clinical study were presented with a series of randomised economic scenarios in order to determine their Willingness To Pay a premium (i.e. an increase) for toric lenses. Then, a similar set of scenarios were presented to a much larger group of online respondents and again, Willingness To Pay was established. Results: For the four measures of visual acuity, the Biofinity toric lens out-performed the Biofinity spherical lens by 0.6 to 1.1 lines.. Subjective vision performance was statistically significantly better with the toric lens for the distance task only. Comfort scores were not significantly different. Similar findings for Willingness To Pay were established for the clinical subjects and for the online respondents. The Willingness To Pay premium (additional fee) for a monthly supply of toric lenses (over spherical lenses) was between £13 and £16, if a toric lens provides better vision and similar comfort, as shown in the clinical study. Conclusion: Consumers are willing to pay a monthly premium of around 50% to benefit from the typical experience of better vision and similar comfort for toric vs. spherical lenses. The level of additional cost for toric lenses compared to their spherical equivalents is less than this in the market, so eye care professionals should consider that toric lenses are delivering a greater clinical return than anticipated by wearers for the relatively small increase in price

Secure and dynamic access control for the Internet of Things (IoT) based traffic system

Today, the trend of the Internet of Things (IoT) is increasing through the use of smart devices, vehicular networks, and household devices with internet-based networks. Specifically, the IoT smart devices and gadgets used in government and military are crucial to operational success. Communication and data sharing between these devices have increased in several ways. Similarly, the threats of information breaches between communication channels have also surged significantly, making data security a challenging task. In this context, access control is an approach that can secure data by restricting unauthorized users. Various access control models exist that can effectively implement access control yet, and there is no single state-of-the-art model that can provide dynamicity, security, ease of administration, and rapid execution all at once. In combating this loophole, we propose a novel secure and dynamic access control (SDAC) model for the IoT networks (smart traffic control and roadside parking management). Our proposed model allows IoT devices to communicate and share information through a secure means by using wired and wireless networks (Cellular Networks or Wi-Fi). The effectiveness and efficiency of the proposed model are demonstrated using mathematical models and discussed with many example implementations.</jats:p

Correlative analyses of RET and RAS mutations in a phase 3 trial of cabozantinib in patients with progressive, metastatic medullary thyroid cancer

BACKGROUND: Cabozantinib significantly prolonged progression-free survival (PFS) versus a placebo in patients with progressive, metastatic medullary thyroid cancer (MTC; P <.001). An exploratory analysis of phase 3 trial data evaluated the influence of rearranged during transfection (RET) and RAS (HRAS, KRAS, and NRAS) mutations on cabozantinib clinical activity. METHODS: Patients (n = 330) were randomized to cabozantinib (140 mg/day) or a placebo. The primary endpoint was PFS. Additional outcome measures included PFS, objective response rates (ORRs), and adverse events in RET and RAS mutation subgroups. RESULTS: Among all study patients, 51.2% were RET mutation–positive (38.2% with RET M918T), 34.8% were RET mutation–unknown, and 13.9% were RET mutation–negative. Sixteen patients were RAS mutation–positive. Cabozantinib appeared to prolong PFS versus the placebo in the RET mutation–positive subgroup (hazard ratio [HR], 0.23; 95% confidence interval [CI], 0.14-0.38; P <.0001), the RET mutation–unknown subgroup (HR, 0.30; 95% CI, 0.16-0.57; P =.0001), and the RAS mutation–positive subgroup (HR, 0.15; 95% CI, 0.02-1.10; P =.0317). The RET M918T subgroup achieved the greatest observed PFS benefit from cabozantinib versus the placebo (HR, 0.15; 95% CI, 0.08-0.28; P <.0001). The ORRs for RET mutation–positive, RET mutation–negative, and RAS mutation–positive patients were 32%, 22%, and 31%, respectively. No PFS benefit was observed in patients lacking both RET and RAS mutations, although the ORR was 21%. The safety profile for all subgroups was similar to that for the overall cabozantinib arm. CONCLUSIONS: These data suggest that cabozantinib provides the greatest clinical benefit to patients with MTC who have RET M918T or RAS mutations. However, a prospective trial is needed to confirm the relation between genetic variation and the response to cabozantinib. Cancer 2016;122:3856–3864. © 2016 American Cancer Society

AP2α controls the dynamic balance between miR-126&126∗ and miR-221&222 during melanoma progression

Accumulating evidences have shown the association between aberrantly expressed microRNAs (miRs) and cancer, where these small regulatory RNAs appear to dictate the cell fate by regulating all the main biological processes. We demonstrated the responsibility of the circuitry connecting the oncomiR-221&222 with the tumor suppressors miR-126&126∗ in melanoma development and progression. According to the inverse correlation between endogenous miR-221&222 and miR-126&126∗, respectively increasing or decreasing with malignancy, their enforced expression or silencing was sufficient for a reciprocal regulation. In line with the opposite roles of these miRs, protein analyses confirmed the reverse expression pattern of miR-126&126∗-targeted genes that were induced by miR-221&222. Looking for a central player in this complex network, we revealed the dual regulation of AP2α, on one side directly targeted by miR-221&222 and on the other a transcriptional activator of miR-126&126∗. We showed the chance of restoring miR-126&126∗ expression in metastatic melanoma to reduce the amount of mature intracellular heparin-binding EGF like growth factor, thus preventing promyelocytic leukemia zinc finger delocalization and maintaining its repression on miR-221&222 promoter. Thus, the low-residual quantity of these two miRs assures the release of AP2α expression, which in turn binds to and induces miR-126&126∗ transcription. All together these results point to an unbalanced ratio functional to melanoma malignancy between these two couples of miRs. During progression this balance gradually moves from miR-126&126∗ toward miR-221&222. This circuitry, besides confirming the central role of AP2α in orchestrating melanoma development and/or progression, further displays the significance of these miRs in cancer and the option of utilizing them for novel therapeutics

Dietary Supplementation with Probiotics Improves Hematopoiesis in Malnourished Mice

BACKGROUND: Lactobacillus rhamnosus CRL1505 (Lr) administered during the repletion of immunocompromised-malnourished mice improves the resistance against intestinal and respiratory infections. This effect is associated with an increase in the number and functionality of immune cells, indicating that Lr could have some influence on myeloid and lymphoid cell production and maturation. OBJECTIVE: This study analyzed the extent of the damage caused by malnutrition on myeloid and lymphoid cell development in the spleen and bone marrow (BM). We also evaluated the impact of immunobiotics on the recovery of hematopoiesis affected in malnourished mice. METHODS: Protein malnourished mice were fed on a balanced conventional diet for 7 or 14 consecutive d with or without supplemental Lr or fermented goat's milk (FGM). Malnourished mice and well-nourished mice were used as controls. Histological and flow cytometry studies were carried out in BM and spleen to study myeloid and lymphoid cells. RESULTS: Malnutrition induced quantitative alterations in spleen B and T cells; however, no alteration was observed in the ability of splenic B cells to produce immunoglobulins after challenge with LPS or CpG. The analysis of BM B cell subsets based on B220, CD24, IgM and IgD expression showed that malnutrition affected B cell development. In addition, BM myeloid cells decreased in malnourished mice. On the contrary, protein deprivation increased BM T cell number. These alterations were reverted with Lr or FGM repletion treatments since normal numbers of BM myeloid, T and B cells were observed in these groups. CONCLUSIONS: Protein malnutrition significantly alters B cell development in BM. The treatment of malnourished mice with L. rhamnosus CRL1505 was able to induce a recovery of B cells that would explain its ability to increase immunity against infections. This work highlights the possibility of using immunobiotics to accelerate the recovery of lymphopoyesis in immunocompromised-malnourished hosts