Antiproliferative mechanism of action of cryptophycin-52: Kinetic stabilization of microtubule dynamics by high-affinity binding to microtubule ends

Cryptophycin-52 (LY355703) is a new synthetic member of the cryptophycin family of antimitotic antitumor agents that is currently undergoing clinical evaluation. At high concentrations (≥10 times the IC₅₀), cryptophycin-52 blocked HeLa cell proliferation at mitosis by depolymerizing spindle microtubules and disrupting chromosome organization. However, low concentrations of cryptophycin-52 inhibited cell proliferation at mitosis (IC₅₀ = 11 pM) without significantly altering spindle microtubule mass or organization. Cryptophycin-52 appears to be the most potent suppressor of microtubule dynamics found thus far. It suppressed the dynamic instability behavior of individual microtubules in vitro (IC₅₀ = 20 nM), reducing the rate and extent of shortening and growing without significantly reducing polymer mass or mean microtubule length. Using [³H]cryptophycin-52, we found that the compound bound to microtubule ends in vitro with high affinity (K_d, 47 nM, maximum of ≈19.5 cryptophycin-52 molecules per microtubule). By analyzing the effects of cryptophycin-52 on dynamics in relation to its binding to microtubules, we determined that ≈5–6 molecules of cryptophycin-52 bound to a microtubule were sufficient to decrease dynamicity by 50%. Cryptophycin-52 became concentrated in cells 730-fold, and the resulting intracellular cryptophycin-52 concentration was similar to that required to stabilize microtubule dynamics in vitro. The data suggest that cryptophycin-52 potently perturbs kinetic events at microtubule ends that are required for microtubule function during mitosis and that it acts by forming a reversible cryptophycin-52-tubulin stabilizing cap at microtubule ends.     

Treatment of haemorrhoids by rubber band ligation.

Outpatient treatment of haemorrhoids by elastic band ligation without anaesthetic was performed in 75 patients. Clinical review showed that 65 returned to work within 24 h of ligation, 8 in 48 h and 2 after 48 h. No patient required hospitalisation for either rectal bleeding or pain after ligation. In the first week after treatment 36 were completely symptom-free, 26 had minimal discomfort and 13 had moderate pain with a sense of imcomplete evacuation in the rectum, and some frequency of call to stool lasting up to 7 days in 1. Follow-up at 8-26 months (mean 10 months) showed that 59 were symptom free, only occasional rectal bleeding occurred in 7 and more frequent irregular bleeding in 2. Religation of residual haemorrhoids was performed in 7 and 2 and a haemorrhoidectomy. For the majority of patients hospitalisation and loss of work were avoided.     

Directing osteogenesis of stem cells with hydroxyapatite precipitated electrospun eri–tasar silk fibroin nanofibrous scaffold

Stimulating stem cell differentiation without growth factor supplement offers a potent and cost-effective scaffold for tissue regeneration. We hypothesise that surface precipitation of nano-hydroxyapatite (nHAp) over blends of non-mulberry silk fibroin with better hydrophilicity and RGD amino acid sequences can direct the stem cell towards osteogenesis. This report focuses on the fabrication of a blended eri–tasar silk fibroin nanofibrous scaffold (ET) followed by nHAp deposition by a surface precipitation (alternate soaking in calcium and phosphate solution) method. Morphology, hydrophilicity, composition, and the thermal and mechanical properties of ET/nHAp were examined by field emission scanning electron microscopy, TEM, FT-IR, X-ray diffraction, TGA and contact angle measurement and compared with ET. The composite scaffold demonstrated improved thermal stability and surface hydrophilicity with an increase in stiffness and elastic modulus (778?±?2.4?N/m and 13.1?±?0.36?MPa) as compared to ET (160.6?±?1.34?N/m and 8.3?±?0.4?MPa). Mineralisation studies revealed an enhanced and more uniform surface deposition of HAp-like crystals, while significant differences in cellular viability and attachment were observed through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and confocal microscopy study. The cell viability and expression of adhesion molecules (CD 44 and CD 29) are found to be optimum for subsequent stages of growth proliferation and differentiation. The rates of proliferation have been observed to decrease owing to the transition of MSC from a state of proliferation to a state of differentiation. The confirmation of improved osteogenic differentiation was finally verified through the alkaline phosphatase assay, pattern of gene expression related to osteogenic differentiation and morphological observations of differentiated cord blood human mesenchymal stem cells under fluorescence microscope. The results obtained showed the improved physicochemical and biological properties of the ET/nHAp scaffold for osteogenic differentiation without the addition of any growth factors.     

Hyponatremia in critically ill patients

Context:

Hyponatremia is a common electrolyte disturbance in critically ill hence understanding its implications is important.

Aims:

This study was carried out to ascertain frequency, predisposing conditions and outcome in critically ill patients with hyponatremia on intensive care unit (ICU) admission.

Settings and Design:

This was an observational, prospective study of a series of ICU patients during a 12-month period.

Materials and Methods:

The patients were divided into two groups: Hyponatremic (serum sodium < 135 mmol/L) and Eunatremic groups (135-145 mmol/L). Clinical examination included volume status and drug history, biochemistries, clinical diagnosis and cause of hyponatremia.

Statistical Analysis Used:

Fisher''s exact test, unpaired t-tests Wilcoxon ranksum tests, profile-likelihood method, log-rank test and Kaplan—Meier curves were used. P < 0.05 were considered to be statistically significant.

Results:

In the hyponatremic group, the frequency of hyponatremia on ICU admission was 34.3%, most were euvolumic, 58.96%. Females comprised of 36.5%. The mean age was 60.4 ± 17.2. The Syndrome of inappropriate Antidiuretic Hormone (SIADH) criteria was met in ninety-one patients (36.25%), peumonia being the leading cause of SIADH. Patients with severe sepsis, elective surgery patients, renal failure and heart failure, cirrhosis of liver and subarachnoid hemorrhage were other more likely etiologic causes (P < 0.05). The hyponatremic group spent a longer time in the ICU (P = 0.02), had longer mechanical ventilator days (P < 0.05) and had an increased mortality rate (P = 0.01).

Conclusions:

Hyponatremia present on admission to the ICU is independent risk factors for poor prognosis.     

Benzophenone assisted UV-activated synthesis of unique Pd-nanodendrite embedded reduced graphene oxide nanocomposite: a catalyst for C–C coupling reaction and fuel cell

In this work we report the use of benzophenone (BP) for the synthesis of a palladium (Pd) embedded on reduced graphene oxide (rGO) nanocomposite (Pd/rGO) using a simple aqueous solution and UV irradiation. The simple and facile evolution of thermodynamically unstable branched Pd(0) nanodendrites was achieved by BP photoactivation, circumventing the growth of more stable nanomorphologies. The synthesis of Pd(0)-embedded rGO nanosheets (PRGO-nd) was made possible by the simultaneous reduction of both the GO scaffold and PdCl₂ by introducing BP into the photoactivation reaction. The nanocomposites obtained in the absence of BP were common triangular and twinned Pd(0) structures which were also implanted on the rGO scaffold (PRGO-nt). The disparity in morphologies presumably occurs due to the difference in the kinetics of the reduction of Pd²⁺ to Pd⁰ in the presence and absence of the BP photoinitiator. It was observed that the PRGO-nd was composed of dense arrays of multiple Pd branches around nucleation site which exhibited (111) facet, whereas PRGO-nt showed a mixture of (100) and (111) facets. On comparing the catalytic efficiencies of the as-synthesized nanocatalysts, we observed a superiority in efficiency of the thermodynamically unstable PRGO-nd nanocomposite. This is due to the evolved active facets of the dendritic Pd(0) morphology with its higher surface area, as testified by Brunauer–Emmett–Teller (BET) analysis. Since both PRGO-nd and PRGO-nt contain particles of similar size, the dents and grooves in the structure are the cause of the increase in the effective surface area in the case of nanodendrites. The unique dendritic morphology of the PRGO-nd nanostructures makes them a promising material for superior catalysis, due to their high surface area, and the high density of surface atoms at their edges, corners, and stepped regions. We investigated the efficiency of the as-prepared PRGO-nd catalyst in the Suzuki–Miyaura coupling reaction and showed its proficiency in a 2 h reaction at 60 °C using 2 mol% catalyst containing 0.06 mol% active Pd. Moreover, the electrochemical efficiency for the catalytic hydrogen evolution reaction (HER) was demonstrated, in which PRGO-nd provided a decreased overpotential of 68 mV for a current density of 10 mA cm⁻², a small Tafel slope of 57 mV dec⁻¹ and commendable stability during chronoamperometric testing for 5 h.

Benzophenone photoinitiator aided synthesis of Pd-nanodendrite embedded rGO nanocatalyst possessing superior potential in C–C coupling reaction and fuel cell application.     

A Survey of Pulmonary Function Abnormalities Following Thoracotomy

Objective

To study the incidence and type of pulmonary function abnormalities after thoracotomy in children.

Methods

Children below 12 y of age who had undergone thoracotomy for any condition and have at least 2 y follow up were included in the study. Detailed assessment of the patients included history and general examination, clinical assessment of pulmonary function, bedside tests to assess pulmonary function and laboratory pulmonary function test using portable spirometer.

Results

Fifty two patients were included in the study. Twenty-seven were cases of esophageal atresia with trachea-esophageal fistula (EATEF), nine pulmonary metastasis from abdominal solid tumors, six mediastinal masses, three hydatid cyst, three eventration of diaphragm, two bronchiectasis, and one each of H-type TEF and congenital esophageal stenosis. The mean age at the time of evaluation was 6.3 y (range 2–18 y). While all the patients were clinically assessed, only 25 (48 %) were eligible for bedside tests and 23 (44 %) for spirometery. The incidences of abnormalities picked were: dyspnea during exercise 8/52 (15.4 %), dyspnea on exercise and on climbing stairs 1/52 (2 %), decreased breath holding time 2/25 (8 %), abnormal incentive spirometry 1/25 (4 %), mild restrictive pattern on pulmonary function test (PFT) 11/23 (47.8 %), moderate restrictive pattern on PFT 2/23 (8.7 %). None had an obstructive pattern on PFT.

Conclusions

Though the incidences of pulmonary function abnormalities were high, these were of mild grade. Close follow up of patients after thoracotomy would be needed for early pick up and appropriate management of these abnormalities to prevent long-term consequences.     

Drilling of bone: A comprehensive review

BackgroundBone fracture treatment usually involves restoring of the fractured parts to their initial position and immobilizing them until the healing takes place. Drilling of bone is common to produce hole for screw insertion to fix the fractured parts for immobilization. Orthopaedic drilling during surgical process causes increase in the bone temperature and forces which can cause osteonecrosis reducing the stability and strength of the fixation.MethodsA comprehensive review of all the relevant investigations carried on bone drilling is conducted. The experimental method used, results obtained and the conclusions made by the various researchers are described and compared.ResultReview suggests that the further improvement in the area of bone drilling is possible. The systematic review identified several consequential factors (drilling parameters and drill specifications) affecting bone drilling on which there no general agreement among investigators or are not adequately evaluated. These factors are highlighted and use of more advanced methods of drilling is accentuated. The use of more precise experimental set up which resembles the actual situation and the development of automated bone drilling system to minimize human error is addressed.ConclusionIn this review, an attempt has been made to systematically organize the research investigations conducted on bone drilling. Methods of treatment of bone fracture, studies on the determination of the threshold for thermal osteonecrosis, studies on the parameters influencing bone drilling and methods of the temperature measurement used are reviewed and the future work for the further improvement of bone drilling process is highlighted.     

Arginine-α, β-dehydrophenylalanine Dipeptide Nanoparticles for pH-Responsive Drug Delivery

Purpose

Nanoparticles (NPs) exhibiting responsiveness towards pH variations in organs, tissue microenvironments and cellular compartments can significantly add on to the drug delivery potential. Here, we have developed NPs from an amphipathic dipeptide, Arginine-α, β-dehydrophenylalanine (RΔF), and tried to explore their pH responsive drug delivery potential in various cancer cells.

Methods

RΔF-NPs were architectured by harnessing the process of molecular self-assembly followed by the assessment of effect of pH on NPs morphology using zetasizer, SEM and CD. FTIR and PXRD analysis of the dipeptide and doxorubicin (Dox) were carried out for compatibility assessment followed by encapsulation of Dox in RΔF-NPs. RΔF-Dox-NPs were evaluated for pH dependent release as well as for in-vitro cellular internalization and efficacy in cancer cells.

Results

RΔF self-assembled to form monodispersed particles at pH 7. SEM analysis revealed a loss of overall particle morphology along with particle aggregation at highly acidic and basic pH respectively. The NPs demonstrated a slow and sustained release behaviour at pH 7 (97.64?±?4.71% after 36 h) in comparison to pH 2 (90.27?±?1.45% after 8 h) and pH 10 (96.39?±?3.87% after 12 h). In-vitro efficacy studies carried-out in various cancer cells revealed that RΔF-Dox-NPs exhibited higher efficacy with 1.65, 1.95 and 13.34 fold lower IC₅₀ values in comparison to Dox in C6, HCT-116 and AGS cell lines.

Conclusions

RΔF-Dox-NPs with higher drug release at acidic pH, enhanced internalization in cancer cells along with higher cytotoxic potential can act as effective pH responsive drug delivery systems.