PLA nanovectors with encapsulated betulin: plant leaf extract-synthesized nanovectors are more efficacious than PVA-synthesized nanovectors

Objectives

Betulin (BT) is an abundant triterpene found predominantly in the bark of Himalayan birch. It is difficult to deliver it in vivo because of its low aqueous solubility. We have therefore developed novel formulations of BT for improving its solubility, bioavailability and therapeutic efficacy.

Results

Poly-d,l-lactide nanovectors (PLA NVs) were synthesized using poly(vinyl alcohol) and Lonicera japonica leaf extract (LE) as a stabiliser and named as PLA-1 NVs and PLA-2 NVs. PLA-1 NVs and PLA-2 NVs were used for the encapsulation of betulin (BT) and named as BT-En-1 and BT-En-2 NVs. The encapsulation efficiency of BT-En-1 and BT-En-2 NVs were 99.3 and 100 % respectively. Prepared nanoformulations were physically stable. An in vitro study revealed 45 % BT was released over 24 h. BT had a prolonged release from BT-En-2 NVs as compared to BT-En-1 NVs. BT-En-2 NVs had better anticancerous activity against SiHa cells than BT-En-1 NVs.

Conclusions

Developed BT-EN-2 NVs had better biocompatibility, excellent stability and enhanced release characteristics than BT-En-1 NVs.

     

Mapping of AFLP markers linked to seed coat colour loci in<Emphasis Type="Italic"> Brassica juncea</Emphasis> (L.) Czern

Association mapping of the seed-coat colour with amplified fragment length polymorphism (AFLP) markers was carried out in 39 Brassica juncea lines. The lines had genetically diverse parentages and varied for seed-coat colour and other morphological characters. Eleven AFLP primer combinations were used to screen the 39 B. juncea lines, and a total of 335 polymorphic bands were detected. The bands were analysed for association with seed-coat colour using multiple regression analysis. This analysis revealed 15 markers associated with seed-coat colour, obtained with eight AFLP primer combinations. The marker E-ACA/M-CTG₃₅₀ explained 69% of the variation in seed-coat colour. This marker along with markers E-AAC/M-CTC₂₃₅ and E-AAC/M-CTA₂₅₀ explained 89% of the total variation. The 15 associated markers were validated for linkage with the seed-coat colour loci using a recombinant inbred line (RIL) mapping population. Bands were amplified with the eight AFLP primer combinations in 54 RIL progenies. Of the 15 associated markers, 11 mapped on two linkage groups. Eight markers were placed on linkage group 1 at a marker density of 6.0 cM, while the remaining three were mapped on linkage group 2 at a marker density of 3.6 cM. Marker E-ACA/M-CTG₃₅₀ co-segregated with Gene1 controlling seed-coat colour; it was specific for yellow seed-coat colour and mapped to linkage group 1. Marker E-AAC/M-CTC₂₃₅ (AFLP8), which had been studied previously, was present on linkage group 2; it was specific for brown seed-coat colour. Since AFLP markers are not adapted for large-scale applications in plant breeding, it is important to convert these to sequence-characterised amplified region (SCAR) markers. Marker E-AAC/M-CTC₂₃₅ (AFLP8) had been previously converted into a SCAR. Work is in progress to convert the second of the linked markers, E-ACA/M-CTG₃₅₀, to a SCAR. The two linked AFLP markers converted to SCARs will be useful for developing yellow-seeded B. juncea lines by means of marker-assisted selection.Communicated by H.F. Linskens     

Life-saving PTMC for critical calcific mitral stenosis in cardiogenic shock with balloon impasse

A 32-year-old male patient, a case of critical calcific mitral stenosis (following closed mitral valvotomy in 1989) was admitted for mitral valve replacement in September 2001. In hospital, he developed cardiogenic shock, pulmonary oedema and oliguria precluding surgery. An emergency percutaneous transatrial balloon mitral commissurotomy as a life-saving procedure in a valve with unfavourable morphology and 'balloon impasse' is discussed.     

Advective hydrogel membrane chromatography for monoclonal antibody purification in bioprocessing

Protein A chromatography is widely employed for the capture and purification of monoclonal antibodies (mAbs). Because of the high cost of protein A resins, there is a significant economic driving force to seek new downstream processing strategies. Membrane chromatography has emerged as a promising alternative to conventional resin based column chromatography. However, to date, the application has been limited to mostly ion exchange flow through (FT) mode. Recently, significant advances in Natrix hydrogel membrane has resulted in increased dynamic binding capacities for proteins, which makes membrane chromatography much more attractive for bind/elute operations. The dominantly advective mass transport property of the hydrogel membrane has also enabled Natrix membrane to be run at faster volumetric flow rates with high dynamic binding capacities. In this work, the potential of using Natrix weak cation exchange membrane as a mAb capture step is assessed. A series of cycle studies was also performed in the pilot scale device (> 30 cycles) with good reproducibility in terms of yield and product purities, suggesting potential for improved manufacturing flexibility and productivity. In addition, anion exchange (AEX) hydrogel membranes were also evaluated with multiple mAb programs in FT mode. Significantly higher binding capacity for impurities (support mAb loads up to 10Kg/L) and 40X faster processing speed were observed compared with traditional AEX column chromatography. A proposed protein A free mAb purification process platform could meet the demand of a downstream purification process with high purity, yield, and throughput. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:974–982, 2015     

Inhibition of galectin-3 mediated cellular interactions by pectic polysaccharides from dietary sources

Pectic polysaccharides from dietary sources such as Decalepis hamiltonii—swallow root (SRPP), Hemidesmus indicus (HPP), Nigella sativa—black cumin (BCPP), Andrographis serpyllifolia—(APP), Zingiber officinale—ginger (GRPP) and, citrus pectin (CPP) were examined for galectin inhibitory activity. Inhibition of (a) galectin-3 of MDA-MB-231 cells induced hemagglutination of red blood cells; (b) galectin-3 mediated interaction between normal/metastatic human buccal cells (NBC)/(MBC) and; (c) invasion of MDA-MB-231 and MBC in the invasive chamber was assessed. Results indicated that SRPP inhibited hemagglutination at Minimum Inhibitory Concentration (MIC) of 1.86 μg ml⁻¹ equivalent of carbohydrate as apposed to those of BCPP (130 μg ml⁻¹), APP (40 μg ml⁻¹), HPP (40 μg ml⁻¹) and CPP (25 μg ml⁻¹). GRPP even at concentration >1–6 mg ml⁻¹ did not inhibit agglutination. Also SRPP showed ∼15 and 2 fold potent anti hemagglutination activity relative to that of galectin-3 specific sugars—galactose (MIC-27.1 μg ml⁻¹) and lactose (MIC-4.16 μg ml⁻¹) respectively. Further, SRPP at 10 μg ml⁻¹ inhibited agglutination of NBC by galectin-3 of MDA-MB-231 cells. Modified swallow root pectic polysaccharide (MSRPP) of 50 kDa retained anti hemagglutination activity (MIC of 1.03 μg ml⁻¹) and inhibited MDA-MB-231 and MBC invasion by 73 and 50% with an IC₅₀ of 136 and 200 μg ml⁻¹ respectively. Both SRPP and MSRPP induced apoptosis up to 80% at 100 μg ml⁻¹ concentration by activating ∼2 and 8 folds of Caspase-3 activity. Sugar composition analysis and its correlation with the galectin inhibitory property indicated that pectic polysaccharides with higher arabinose and galactose content—arabinogalactan inhibited hemagglutination significantly.     

ABL/BCR gene variant with two-step mechanism: Unusual localization and rare/novel chromosomal rearrangements in CML patients

Aims: Variant translocations involving 9q, 22q and at least one additional genomic locus occur in 5-10% of the patients with chronic myeloid leukemia (CML). The mechanisms for the formation of these variant translocations are not fully characterized. Here we report CML cases presenting a variant translocation indicating two-step mechanism with rare/novel chromosomal rearrangement. Methods: Karyotype analysis was performed on metaphases obtained through short-term cultures of bone marrow and blood. Detection of BCR-ABL fusion gene was performed using dual-color dual-fusion (D-FISH) and extra signal (ES) translocation probes. BAC-FISH was also carried out. Results: In Patient 1, the third partner chromosome was der(11)(p15) with a 2F2G1R signal pattern, which is an unusual signal pattern with the two-step mechanism. Patients 2 and 3 showed typical positive (2F1G1R) signal pattern. In Patient 2, both the chromosome 22s were involved in variant formation. The second fusion was observed below the BCR gene of the second homologue. In Patient 3 the third chromosome was der(13)(q14). The fourth patient showed a variant pattern with BCR/ABL-ES probe involving der(X)(q13) region. Conclusion: The presence of different rearrangements of both 9q34 and 22q11 regions highlights the genetic heterogeneity of this subgroup of CML. In each case with variants, further studies with FISH, BAC-FISH or more advanced technique such as microarray should be performed. Future studies should be performed to confirm the presence of true breakpoint hot spots and assess their implications in CML with variant Ph.     

A Hump-Backed Trend in Bacterial Diversity with Elevation on Mount Fuji, Japan

Little is known of how bacterial diversity in soils varies with elevation. One previous study found a decline with elevation, whereas another found no trend. We chose Mount Fuji of Japan as a geologically and topographically simple mountain system. Samples were taken at elevational intervals, between the base of the mountain at 1,000 m and its summit at 3,700 m. Polymerase chain reaction-amplified soil DNA for the bacterial 16S gene targeting V1–V3 region was pyrosequenced using the 454 Roche machine, and taxonomically classified with reference to a bioinformatic database. There was a significant “peak” in total bacterial diversity at around 2,500 m above the tree line with a decline towards the highest elevations around 3,700 m near the summit. Individual bacterial phyla show distinct trends—increase, decrease, or a mid-elevational “bulge” in diversity. Bacterial diversity does not parallel woody plant or herbaceous plant diversity. We suggest that beyond the tree and vegetation line, the more extreme temperature fluctuations, stronger UV, lack of nutrients, and more frequent disturbance of the loose substrate of these slopes allows less competition and greater bacterial species diversity due to “lottery” recruitment. However, at the highest elevations, the physiological challenges are so extreme that fewer bacterial species are capable of surviving.     

Pelvic Floor Dysfunction in Morbidly Obese Women: Pilot Study

Objective: The objective of this study was to evaluate the impact of obesity on pelvic floor function in women. Research Methods and Procedures: This was a prospective controlled study of 20 morbidly obese female patients planning to undergo gastric bypass surgery and 20 age‐matched female controls. Subjects completed symptom and impact questionnaires, including the Incontinence Impact Questionnaire (IIQ‐7), Urogenital Distress Inventory (UDI), the Kobashi Prolapse Symptom Inventory and Quality‐of‐Life Questionnaire (PSI‐QOL), and Index of Female Sexual Function. Data were analyzed with Wilcoxon or ratio χ² tests. Results: Mean weight was 295.7 ± 87.9 lbs in the study group and 144.79 ± 33.07 lbs in the control group. Mean BMI was 52.65 ±14.49 kg/m² in the study group and 25.11 ± 5.27 kg/m² in the control group. According to the IIQ‐7, urinary incontinence significantly affected lifestyle in the study group. The total IIQ‐7 score was also significantly affected in the study group (p = 0.03). The UDI indicated more urinary leakage with activity (p = 0.04) and more incidents of small amounts of leakage (p = 0.02) in the study group. According to the PSI‐QOL, women in the study group experienced constipation more often because of difficulty in emptying the rectum (p = 0.04). The PSI‐QOL score was higher in the study group (6.75 ± 6.84) than in the control group (2.65 ± 3.03; p = 0.04). There were no significant differences between groups regarding sexual function. Discussion: Morbid obesity is associated with a significant negative impact on urogenital health. Sexual function did not seem to be affected in women who are morbidly obese.