Scaffolds for drug delivery and tissue engineering: The role of genetics.

Scaffolds are implants commonly used to deliver cells, drugs, and genes into the body. Their regular porous structure ensures the proper support for cell attachment, proliferation, differentiated function, and migration. Techniques to fabricate a scaffold include leaching, freeze-drying, supercritical fluid technology, thermally induced phase separation, rapid prototyping, powder compaction, sol-gel, and melt molding. Gene delivery from the scaffold represents a versatile approach to influence the environment for managing cell function. Scaffolds can be used for various tissue engineering purposes, e.g. bone formation, periodontal regeneration, cartilage development, artificial corneas, heart valves, tendon repair, or ligament replacement. Moreover, they are also instrumental in cancer therapy, inflammation, diabetes, heart disease, and wound dressings. Scaffolds provide a platform to extend the delivery of drugs and genetic materials at a controlled timeframe, besides potentially being used to prevent infection upon surgery and other chronic diseases, provided that they can be formulated with specific medicines. This review discusses the need to design advanced functional scaffolds with the potential for modified drug delivery and tissue engineering in a synergistic approach. Special attention is given to works published in 2023 to generate the bibliometric map.

THC-Reduced Cannabis sativa L.-How Does the Solvent Determine the Bioavailability of Cannabinoids Given Orally?

The bioavailability levels of cannabidiol (CBD) and tetrahydrocannabinol (THC) determine their pharmacological effects. Therefore, for medical purposes, it is essential to obtain extracts containing the lowest possible content of the psychogenic component THC. In our extract, the CBD/THC ratio was 16:1, which is a high level compared to available medical preparations, where it is, on average, 1:1. This study assessed the bioavailability and stability of CBD and THC derived from Cannabis sativa L. with reduced THC content. The extract was orally administered (30 mg/kg) in two solvents, Rapae oleum and Cremophor, to forty-eight Wistar rats. The whole-blood and brain concentrations of CBD and THC were measured using liquid chromatography coupled with mass spectrometry detection. Much higher concentrations of CBD than THC were observed for both solvents in the whole-blood and brain after oral administration of the Cannabis sativa extract with a decreased THC content. The total bioavailability of both CBD and THC was higher for Rapae oleum compared to Cremophor. Some of the CBD was converted into THC in the body, which should be considered when using Cannabis sativa for medical purposes. The THC-reduced hemp extract in this study is a promising candidate for medical applications.

Tocilizumab-coated solid lipid nanoparticles loaded with cannabidiol as a novel drug delivery strategy for treating COVID-19: A review.

Commonly used clinical strategies against coronavirus disease 19 (COVID-19), including the potential role of monoclonal antibodies for site-specific targeted drug delivery, are discussed here. Solid lipid nanoparticles (SLN) tailored with tocilizumab (TCZ) and loading cannabidiol (CBD) are proposed for the treatment of COVID-19 by oral route. TCZ, as a humanized IgG1 monoclonal antibody and an interleukin-6 (IL-6) receptor agonist, can attenuate cytokine storm in patients infected with SARS-CoV-2. CBD (an anti-inflammatory cannabinoid and TCZ agonist) alleviates anxiety, schizophrenia, and depression. CBD, obtained from Cannabis sativa L., is known to modulate gene expression and inflammation and also shows anti-cancer and anti-inflammatory properties. It has also been recognized to modulate angiotensin-converting enzyme II (ACE2) expression in SARS-CoV-2 target tissues. It has already been proven that immunosuppressive drugs targeting the IL-6 receptor may ameliorate lethal inflammatory responses in COVID-19 patients. TCZ, as an immunosuppressive drug, is mainly used to treat rheumatoid arthritis, although several attempts have been made to use it in the active hyperinflammatory phase of COVID-19, with promising outcomes. TCZ is currently administered intravenously. It this review, we discuss the potential advances on the use of SLN for oral administration of TCZ-tailored CBD-loaded SLN, as an innovative platform for managing SARS-CoV-2 and related infections.

Simple Extraction of Cannabinoids from Female Inflorescences of Hemp (Cannabis sativa L.).

The high interest in non-psychoactive cannabidiol increases the need for efficient and straightforward cannabidiol (CBD) extraction methods. The research aimed to compare simple methods of cannabinoid extraction that do not require advanced laboratory equipment. This work assesses the content of total CBD and Δ9-tetrahydrocannabinol (Δ9-THC) in popular solvents such as water and ethanol extracts. Hemp raw material was analyzed with Gas Chromatography with a Flame Ionization Detector (GC-FID), while extracts were tested by High-Performance Liquid Chromatography (HPLC). The female inflorescences of three varieties of industrial hemp were tested: Futura 75, KC Dora, and Tygra (different sowing and N fertilization densities). Tygra (T/10/30) showed the highest content of CBD (0.064%) in water extracts. However, in 80% tincture from Futura 75 (F/30/30), a higher CBD content of 1.393% was observed. The use of 96% ethanol for extraction and ultrasound enabled the highest CBD content to be obtained: 2.682% in Futura 75 (F/30/30). Cold water extraction showed no effect on Δ9-THC content, while hot water extraction increased content from 0.001% in KC Dora to 0.002% in Futura 75 (F/30/30) and Tygra, but the changes were statistically insignificant. Application of 80% ethanol revealed the significantly highest content of Δ9-THC in KC Dora, from 0.026% (K/30/90) to 0.057% (K/30/30), as well as in Tygra (T/30/30) (0.036%) and Futura 75 (F/30/30) (0.048%). The use of ethanol extraction in combination with ultrasound could be an efficient method of obtaining cannabinoids.

Lipid-Drug Conjugates and Nanoparticles for the Cutaneous Delivery of Cannabidiol.

Lipid nanoparticles are currently used to deliver drugs to specific sites in the body, known as targeted therapy. Conjugates of lipids and drugs to produce drug-enriched phospholipid micelles have been proposed to increase the lipophilic character of drugs to overcome biological barriers. However, their applicability at the topical level is still minimal. Phospholipid micelles are amphiphilic colloidal systems of nanometric dimensions, composed of a lipophilic nucleus and a hydrophilic outer surface. They are currently used successfully as pharmaceutical vehicles for poorly water-soluble drugs. These micelles have high in vitro and in vivo stability and high biocompatibility. This review discusses the use of lipid-drug conjugates as biocompatible carriers for cutaneous application. This work provides a metadata analysis of publications concerning the conjugation of cannabidiol with lipids as a suitable approach and as a new delivery system for this drug.

Cancer Nanopharmaceuticals: Physicochemical Characterization and In Vitro/In Vivo Applications.

Physicochemical, pharmacokinetic, and biopharmaceutical characterization tools play a key role in the assessment of nanopharmaceuticals' potential imaging analysis and for site-specific delivery of anti-cancers to neoplastic cells/tissues. If diagnostic tools and therapeutic approaches are combined in one single nanoparticle, a new platform called nanotheragnostics is generated. Several analytical technologies allow us to characterize nanopharmaceuticals and nanoparticles and their properties so that they can be properly used in cancer therapy. This paper describes the role of multifunctional nanoparticles in cancer diagnosis and treatment, describing how nanotheragnostics can be useful in modern chemotherapy, and finally, the challenges associated with the commercialization of nanoparticles for cancer therapy.

Two- and Three-Dimensional Spectrofluorimetric Qualitative Analysis of Selected Vegetable Oils for Biomedical Applications.

Vegetable oils obtained from different plants are known for their beneficial effects on prophylaxis and supportive treatment of a great deal of inflammatory-mediated conditions. Their wide range of saturated and unsaturated fatty acids, and the presence of other ingredients (e.g., tocopherols, chlorophylls), provide them with anti-inflammatory, antioxidant and anticancer properties, which are worth being exploited. In this study, we have carried out the spectrofluorometric analysis of selected vegetable oils, namely apricot (Prunus armeniaca) kernel oil; blueberry (Vaccinium spp.) seed oil; argan (Argania spinosa) nut oil; kiwi (Actinidia deliciosa) seed oil; grape (Vitis vinifera) seed oil; evening primrose (Oenothera biennis) oil and meadowfoam (Limnanthes alba) seed oil, with the purpose to detect their fluorescent ingredients for further identification and bioactivity comparison. The obtained two- (2D) and three-dimensional (3D) emission spectra offered a complete description of the fluorescent components of the mixture and revealed different features for studied oils.

Is Polymorphism in the Apoptosis and Inflammatory Pathway Genes Associated With a Primary Response to Anti-TNF Therapy in Crohn's Disease Patients?

Anti-tumor necrosis factor (TNF) therapy is used for the induction and maintenance of remission in Crohn's disease (CD) patients. However, primary nonresponders to initial treatment constitute 20%-40% of cases. The causes of this phenomenon are still unknown. In this study, we aimed to determine the genetic predictors of the variable reactions of CD patients to anti-TNF therapy. Using long-range PCR libraries and the next-generation sequencing (NGS) method, we performed broad pharmacogenetic studies including a panel of 23 genes (TNFRSF1A, TNFRSF1B, CASP9, FCGR3A, LTA, TNF, FAS, ADAM17, IL17A, IL6, MMP1, MMP3, S100A8, S100A9, S100A12, TLR2, TLR4, TLR9, CD14, IL23R, IL23, IL1R, and IL1B) in a group of 107 diagnosed and clinically characterized CD patients following anti-TNF therapy. In the studied group, we indicated, in total, 598 single nucleotide variants for all analyzed genomic targets. Twelve patients (11.2%) did not respond to the induction therapy, which was associated with alleles in 11 loci located in FCGR3A (rs7539036, rs6672453, rs373184583, and rs12128686), IL1R (rs2041747), TNFRSF1B (rs5746053), IL1B (rs1071676, rs1143639, rs1143637, and rs1143634), and FAS (rs7896789) genes. After multiple comparison corrections, the results were not statistically significant, however for nonresponders the alleles distribution for those loci presented large differences and specified scheme compared to responders and populations. These findings require further investigation in an independent larger cohort before introducing them for a clinical setting, however, we identified an interesting direction. Polymorphism of the FCGR3A, IL1R, TNFRSF1B, IL1B, and FAS genes could be a predictor of the primary response to anti-TNF therapy in CD patients.

Transgenic Plant Detection Using an AuNPs Based SPR Biosensor.

The intensive development and commercialization of genetically modified plants observed over the last decade has led to the development of transgenic detection methods that are rapid and sensitive. Among the strategies used for the detection/monitoring of genetically modified organisms (GMOs), surface plasmon resonance (SPR) meets the necessary criteria. This optical technique measures the changes in the refractive index in the vicinity of thin metal layers (i.e., gold) in response to biomolecular interactions occurring at a flat metal‒solution interface. Additionally, it allows the application of functionalized gold nanoparticles (AuNPs) in SPR research to enhance the signal intensity. In the present study, an SPR method, enhanced by the application of AuNPs, was developed to detect transgenic tobacco plants carrying a Streptococcus mutans antigen. The basis for the detection of the target DNA was the hybridization between the genomic DNA isolated from the leaves, stems, and roots of the transgenic tobacco and the biotinylated oligonucleotide probes immobilized onto a streptavidin (SA) sensor chip. SA-functionalized AuNPs coated with a second type of biotinylated probe were applied to increase the sensitivity of the detection method. Analysis of the results indicated that the constructed SPR-based sensor chip can potentially recognize complementary standard fragments (nonamplified genomic DNA) at concentrations as low as 1 pM. Thus, nonamplified transgenic DNA was detected using a label-free and real-time AuNPs-enhanced SPR biosensing method. This unique approach could be used to detect GMOs with high efficiency, even at a low detection limit, high repeatability, and with less time and a lower cost needed for each analysis.

ESR1 Gene Variants Are Predictive of Osteoporosis in Female Patients with Crohn's Disease.

Decreased bone mass in patients with inflammatory bowel diseases (IBD) is a clinical problem with extremely severe consequences of osteoporotic fractures. Despite its increasing prevalence and the need for mandatory intervention and monitoring, it is often ignored in IBD patients' care. Determining the biomarkers of susceptibility to bone mineral density disorder in IBD patients appears to be indispensable. We aim to investigate the impact of estrogen receptor gene (ESR1) gene polymorphisms on bone mineral density (BMD) in patients with ulcerative colitis (UC) and Crohn's disease (CD), as they may contribute both, to osteoporosis and inflammatory processes. We characterised 197 patients with IBD (97 with UC, 100 with CD), and 41 controls carrying out vitamin D, calcium and phosphorus serum levels, and bone mineral density assessment at the lumbar spine and the femoral neck by dual-energy X-ray absorptiometry (DXA), ESR1 genotyping and haplotype analysis. We observed that women with CD showed the lowest bone density parameters, which corresponded to the ESR1 c.454-397T and c.454-351A allele dose. The ESR1 gene PvuII and XbaI TA (px) haplotype correlated with decreased femoral neck T-score (OR = 2.75, CI = [1.21-6.27], P-value = 0.016) and may be predictive of osteoporosis in female patients with CD.

Vitamin D receptor (VDR) TaqI polymorphism, vitamin D and bone mineral density in patients with inflammatory bowel diseases.

BACKGROUND: A common feature in the etiology of inflammatory bowel disease (IBD) and osteoporosis is a complex genetic background. Moreover, it has been shown that some of the susceptibility loci overlap for both diseases. One of the genes that may be involved in the pathogenesis of IBD as well as decreased bone mass is the vitamin D receptor (VDR) gene. OBJECTIVES: The aim of this study was to investigate the association of the TaqI polymorphism (rs731236, c.1056T >C) in the VDR gene with serum vitamin D concentration and bone mineral density (BMD) in patients with IBD. MATERIAL AND METHODS: A total of 172 IBD patients (85 with Crohn's disease (CD) and 87 with ulcerative colitis (UC)) and 39 healthy controls were enrolled in the study. Polymorphism was determined with polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). Bone mineral density was measured at the lumbar spine (L2-L4) and the femoral neck (FN) using dual-energy x-ray absorptiometry (DEXA). Serum concentrations of 25-hydroxyvitamin D were determined using electrochemiluminescence binding assay (ECLIA). RESULTS: Our studies revealed that serum vitamin D concentration in IBD patients was not lowered in comparison with healthy controls. Patients with CD presented more advanced osteopenia and osteoporosis. Individuals with UC carrying the TaqI tt genotype of VDR gene showed significantly higher FN BMD than carriers of TT and Tt genotypes (p = 0.02). Moreover, tt genotype was present with higher frequency in UC patients than in controls and CD patients (23% vs 7.7% and 16.5%, respectively). CONCLUSIONS: The tt genotype may have a protective effect on BMD in UC patients.

DNA variants in Helicobacter pylori infected patients with chronic gastritis, dysplasia and gastric cancer.

PURPOSE: The main scope of this study was to evaluate the importance of selected DNA variants for developing inflammation of gastric mucosa and carcinogenesis in gastrointestinal diseases in patients infected with Helicobacter pylori. PATIENTS AND METHODS: Patients subjected to analysis constituted a group of 131 consecutive cases, with control groups consisting of 100 healthy volunteers and 13 dyspeptic patients. Molecular analysis included the following genes: TP53 (c.743 G > A, c.746 G > A, c.749C > T), MSH2 (c.942 + 3A > T), MLH1 (c.2041 G > A), NOD2/CARD15 (c.3016_3017insC, c.802C > T), IL1A (c.-949C > T) and IL1B (c.315C > T). DNA variants were detected using PCR-RFLP, pyrosequencing and sequencing. RESULTS: Mutations of the analyzed genes were observed more frequently in patients with a higher degree of mucosal lesions (50.9%) than in patients with milder mucosal changes (27.6%). Single mutations and polymorphisms did not affect the course of the disease. Our analysis confirms the influence of the NOD2/CARD15 c.802C > T polymorphism on the development of mucosal changes. A correlation of the frequency of the CT genotype of the NOD2/CARD15 c.802C > T polymorphism with the NOD2/CARD15 c.3016_3017insC mutation was observed. The TT genotype frequency in the c.315C > T IL1B gene polymorphism was statistically significantly higher in patients with mucosa changes. CONCLUSIONS: Accumulation of molecular abnormalities may increase the susceptibility to inflammatory response of the gastric mucosa in H. pylori-infected patients and play an important role in the development of chronic active gastritis, atrophy, intestinal metaplasia, dysplasia and the intestinal type of gastric cancer. The severity of gastric mucosal damage correlates with the presence of mutations in the gastric mucosa and the age of patients.

Modifying impact of RET gene haplotypes on medullary thyroid carcinoma clinical course.

The clinical course of medullary thyroid carcinoma (MTC) associated with the MEN2A syndrome as well as of sporadic MTC shows considerable heterogeneity. The disease picture varies not only between the same RET proto-oncogene mutation carriers but also among sporadic MTC patients with no RET germinal mutations, which suggests the involvement of additional modulators of the disease. However, genetic factors responsible for this heterogeneity of the MTC clinical course still remain unknown. The aim of this study was to determine if polymorphic variants or specific haplotypes of the RET gene may modify the MTC clinical course. We genotyped the following loci: c.73+9277T>C, c.135G>A, c.1296A>G, c.2071G>A, c.2307T>C, c.2508C>T and c.2712C>G in 142 MTC patients and controls. We demonstrated considerable differences in the genotypes distribution within c.73+9277T>C, c.135G>A and c.2307T>C loci Our results show that the c.73+9277T variant associated with a decreased activity of the MCS+9.7 RET enhancer is rare in hereditary MTC patients with primary hyperparathyroidism, and thus, may influence the MTC clinical picture. The decreased activity of the RET promoter enhancer reduces RET expression level and may counterbalance the activating mutation in this gene. Frequent co-occurrence of the c.73+9277T allele with p.E768D, p.Y791F, p.V804M or p.R844Q RET mutations may be associated with their attenuation and milder clinical picture of the disease. Haplotypes analysis showed that C-G-A-G-T-(C)-C (c.73+9277T>C - c.135G>A - c.1296A>G - c.2071G>A - c.2307T>G - (c.2508C>T) - c.2712C>G) alleles combination predisposes to pheochromocytomas and primary hyperparathyroidism. We consider that RET haplotypes defining may become an auxiliary diagnostic tool in MTC patients.

Longrange PCR-based next-generation sequencing in pharmacokinetics and pharmacodynamics study of propofol among patients under general anaesthesia.

The individual response of patients to propofol results from the influence of genetic factors. However, the state of knowledge in this matter still remains insufficient. The aim of our study was to determine genetic predictors of variable pharmacokinetics and pharmacodynamics of propofol within selected 9 genes coding for propofol biotransformation enzymes, receptors and transporters. Our studies are the first extensive pharmaocgenetics research of propofol using high throughput sequencing technology. After the design and optimization of long range PCR-based next-generation sequencing experiment, we screened promoter and coding sequences of all genes analyzed among 87 Polish patients undergoing general anaesthesia with propofol. Initially we found that two variants, c.516 G > T in the CYP2B6 gene and c.2677 T > G in the ABCB1 gene, significantly correlate with propofol's metabolic profile, however after Bonferroni correction the P-values were not statistically significant. Our results suggest, that variants within the CYP2B6 and ABCB1 genes correlate stronger with propofol's metabolic profile compared to other 7 genes. CYP2B6 and ABCB1 variants can play a potentially important role in response to this anaesthetic and they are promising object for further studies.

Human gingival fibroblast response to enamel matrix derivative, porcine recombinant 21.3-kDa amelogenin and 5.3-kDa tyrosine-rich amelogenin peptide.

Enamel matrix derivative (EMD) containing a variety of protein fractions has been used for periodontal tissue regeneration. It is suggested that the proteins contained in EMD positively influence gingival fibroblasts migration and proliferation. Effects of EMD as well as of porcine recombinated 21.3-kDa amelogenin (prAMEL) and 5.3-kDa tyrosine-rich amelogenin peptide (prTRAP) on human gingival fibroblast (HGF-1, ATCC; USA) cell line were investigated. Real-time cell analysis (xCELLigence system; Roche Applied Science) was performed to determine the effects of EMD, prAMEL and prTRAP (12.5-50 µg/mL) on HGF-1 cell proliferation and migration. The effect of treatment on cell cycle was determined using flow cytometry. EMD significantly increased HGF-1 cell proliferation after 24- and 48-h incubation. Individually, prAMEL and prTRAP also increased HGF-1 cell proliferation; however, the difference was significant only for prAMEL 50 µg/mL. prAMEL and TRAP significantly increased HGF-1 cell migration after 60- and 72-h incubation. Cell cycle analysis showed significant decrease of the percentage of cells in the G0/G1 phase and a buildup of cells in the S and M phase observed after EMD and prAMEL stimulation. This process was ligand and concentration-dependent. The various molecular components in the enamel matrix derivative might contribute to the reported effects on gingival tissue regeneration; however, biologic effects of prAMEL and prTRAP individually were different from that of EMD.

Effects of enamel matrix proteins on adherence, proliferation and migration of epithelial cells: A real-time in vitro study.

Enamel matrix derivative (EMD) can mimic odontogenic effects by inducing the proliferation and differentiation of connective tissue progenitor cells, stimulating bone growth and arresting epithelial cells migration. To the best of our knowledge, there is no data indicating that any active component of EMD reduces epithelial cell viability. The present study examines the impact of commercial lyophilized EMD, porcine recombinant amelogenin (prAMEL; 21.3 kDa) and tyrosine-rich amelogenin peptide (TRAP) on the adherence, proliferation and migration of human epithelial cells in real-time. The tongue carcinoma cell line SCC-25 was stimulated with EMD, porcine recombinant AMEL and TRAP, at concentrations of 12.5, 25 and 50 µg/ml. Cell adherence, migration and proliferation were monitored in real-time using the xCELLigence system. No significant effects of EMD on the morphology, adhesion, proliferation and migration of SCC-25 cells were observed. However, porcine recombinant AMEL had a dose-dependent inhibitory effect on SCC-25 cell proliferation and migration. Predominantly, no notable differences were found between control and TRAP-treated cells in terms of cell adhesion and migration, a decrease in proliferation was observed, but this was not statistically significant. EMD and its active components do not increase the tongue cancer cell viability.

Effects of Intrauterine Environment on the Magnitude of Differences Within the Pairs of Monozygotic and Dizygotic Twins.

The aim of this study was to determine the effects of intrauterine environment on the magnitude of intrapair differences in six somatic traits of monozygotic (MZ) and dizygotic (DZ) twins (1,263 pairs; 424 MZ twins and 839 DZ twins). Differences in intrauterine environments of MZ twins enforced division of the research material into four groups: (1) MZ-MC-TTTS - MZ twins from monochorionic (MC) pregnancies with twin-to-twin transfusion syndrome (TTTS), (2) MZ-MC (without TTTS)-MZ twins from MC pregnancies without TTTS, (3) MZ-DC-MZ twins from dichorionic (DC) pregnancies, and (4) DZ-DZ twins. The intrapair differences in all analyzed somatic traits, especially body weight and circumference of the chest, were the largest in the case of MZ twins from MC pregnancies with TTTS. DZ twins were the group presenting with the second largest intrapair differences in the analyzed traits. At the end of pregnancy, that is, in lunar months 9 and 10, the magnitude of intrapair differences in all traits of twins from this group was significantly greater than in MZ twins from both MC and DC pregnancies. Irrespective of the analyzed period, the least evident, statistically insignificant intrapair differences in the studied traits were documented in the case of MZ twins from MC pregnancies without TTTS and twins from DC pregnancies. These findings imply that the differentiating effect of intrauterine environment is associated with the occurrence of TTTS, rather than with chorionicity, as postulated previously.

Impact of CYP2E1, GSTA1 and GSTP1 gene variants on serum alpha glutathione S-transferase level in patients undergoing anaesthesia.

BACKGROUND: The serum glutathione S-transferase alpha (α-GST) concentration has been used as a marker of hepatic condition. After sevoflurane anaesthesia a mild impairment of hepatocellular integrity was observed. Genetic polymorphisms in CYP2E1, GSTA1 and GSTP1 genes, affecting enzymes activity, may possibly influence the hepatotoxic effect of sevoflurane. The aim of this study was to assess the influence of genetic polymorphism of CYP2E1, GSTA1 and GSTP1 genes on serum α-GST level in 86 unrelated patients representing ASA physical status I-II, undergoing laryngological surgery under general anaesthesia with sevoflurane. METHODS: The serum samples from three perioperative time points were analyzed using ELISA. Genetic variants were detected by pyrosequencing and sequencing. Finally, the statistical associations between serum α-GST concentration and analyzed alleles of CYP2E1, GSTP1 and GSTA1 genes were estimated. RESULTS: The allele GSTA1*B (-567G, -69T, -52A) frequency was 0.43, whereas the alleles c.313G and c.341T of GSTP1 were identified with frequencies of 0.28 and 0.1 respectively. The -1053T allele of the CYP2E1 gene was observed with 0.01 frequency. We found serum α-GST concentrations in homozygous changes c.313A>G and c.341C>T of the GSTP1 gene significantly higher at the end of anaesthesia as compared with the levels at pre-anaesthetic and 24 h post-anaesthetic time points. Moreover, GSTA1 wild type genotype was associated with increased α-GST concentration at 24 h after the end of anaesthesia. CONCLUSIONS: GSTP1 gene polymorphism has an impact on the perioperative serum α-GST concentration in patients undergoing sevoflurane anaesthesia. A similar association, although not statistically significant exists between GSTA1 gene variants and perioperative serum α-GST level.

Ecosensitivity and genetic polymorphism of somatic traits in the perinatal development of twins.

In view of criticism regarding the usefulness of heritability coefficients, the aim of this study was to analyze separately the information on genetic and environmental variability. Such an approach, based on the normalization of trait's variability for its value, is determined by the coefficients of genetic polymorphism (Pg) and ecosensitivity (De). The studied material included 1263 twin pairs of both sexes (among them 424 pairs of monozygotic twins and 839 pairs of dizygotic twins) born between the 22nd and 41st week of gestation. Variability of six somatic traits was analyzed. The zygosity of same-sex twins was determined based on the polymorphism of DNA from lymphocytes of the umbilical cord blood, obtained at birth. The coefficients of genetic polymorphism and ecosensitivity for analyzed traits of male and female twins born at various months of gestation were calculated. Our study revealed that a contribution of the genetic component predominated over that of the environmental component in determining the phenotypic variability of somatic traits of newborns from twin pregnancies. The genetically determined phenotypic variability in male twins was greater than in the females. The genetic polymorphism and ecosensitivity of somatic traits were relatively stable during the period of fetal ontogeny analyzed in this study. Only in the case of body weight, a slight increase in the genetic contribution of polygenes to the phenotypic variance could be observed with gestational age, along with a slight decrease in the influence of environmental factors.

1,3-Propanediol production by Escherichia coli using genes from Citrobacter freundii atcc 8090.

Compared with chemical synthesis, fermentation has the advantage of mass production at low cost, and has been used in the production of various industrial chemicals. As a valuable organic compound, 1,3-propanediol (1,3-PDO) has numerous applications in the production of polymers, lubricants, cosmetics and medicines. Here, conversion of glycerol (a renewable substrate and waste from biodiesel production) to 1,3-PDO by E. coli bacterial strain carrying altered glycerol metabolic pathway was investigated. Two gene constructs containing the 1,3-PDO operon from Citrobacter freundii (pCF1 and pCF2) were used to transform the bacteria. The pCF1 gene expression construct contained dhaBCE genes encoding the three subunits of glycerol dehydratase, dhaF encoding the large subunit of the glycerol dehydratase reactivation factor and dhaG encoding the small subunit of the glycerol dehydratase reactivating factor. The pCF2 gene expression construct contained the dhaT gene encoding the 1,3-propanediol dehydrogenase. Expression of the genes cloned in the above constructs was under regulation of the T7lac promoter. RT-PCR, SDS-PAGE analyses and functional tests confirmed that 1,3-PDO synthesis pathway genes were expressed at the RNA and protein levels, and worked flawlessly in the heterologous host. In a batch flask culture, in a short time applied just to identify the 1,3-PDO in a preliminary study, the recombinant E. coli bacteria produced 1.53 g/L of 1,3-PDO, using 21.2 g/L of glycerol in 72 h. In the Sartorius Biostat B Plus reactor, they produced 11.7 g/L of 1,3-PDO using 24.2 g/L of glycerol, attaining an efficiency of 0.58 [mol1,3-PDO/molglycerol].