Bacteriological profile and frequency of antibiotic resistance in the infected diabetic foot.

The diabetic foot (DF) is a major public health problem, with infection of the DF being a major risk factor for amputation. This study aimed to establish the bacteriological profile of diabetic foot infection and to evaluate the resistance of isolated bacteria to antibiotics. Retrospective study of 150 patients in the internal medicine departments of the University Hospital of Batna, the University Hospital of Constantine and the EPH of Oum El Bouaghi, over 24 months from September 2019 to August 2021. The population had an average age of 46±13 years and a sex ratio of 1.88 in favour of men. The majority of patients were type 2 diabetics with a mean duration of the evolution of 20 years. The starting point of the foot lesions was inter-toe intertrigo and trauma. The most frequent lesion found was an infectious lesion followed by gangrene, classified in 33.64% of cases as Grade 2 according to Wagner. We performed a swab of the ulcer, and direct examination showed a predominance of Gram-negative bacilli (GNB) in 78.56% of the samples, 17 different bacterial species were isolated, predominantly Escherichia coli and Pseudomenas aeruginosa. We studied the antibiotic resistance of the most frequently isolated bacteria, the antibiotic therapy was initially probabilistic and then adapted to the antibiogram. The multidisciplinary management of diabetic foot is still very precarious, and the best treatment is prevention.

Characterization and Pharmacokinetic Evaluation of Oxaliplatin Long-Circulating Liposomes.

The clinical efficacy of Oxaliplatin (L-OHP) is potentially limited by dose-dependent neurotoxicity and high partitioning to erythrocytes in vivo. Long-circulating liposomes could improve the pharmacokinetic profile of L-OHP and thus enhance its therapeutic efficacy and reduce its toxicity. The purpose of this study was to prepare L-OHP long-circulating liposomes (L-OHP PEG lip) by reverse-phase evaporation method (REV) and investigate their pharmacokinetic behavior based on total platinum in rat plasma using atomic absorption spectrometry (AAS). A simple and a sensitive AAS method was developed and validated to determine the total platinum originated from L-OHP liposomes in plasma. Furthermore, long-circulating liposomes were fully characterized in vitro and showed great stability when stored at 4°C for one month. The results showed that the total platinum in plasma of L-OHP long-circulating liposomes displayed a biexponential pharmacokinetic profile with five folds higher bioavailability and longer distribution half-life compared to L-OHP solution. Thus, long-circulating liposomes prolonged L-OHP circulation time and may present a potential candidate for its tumor delivery. Conclusively, the developed AAS method could serve as a reference to investigate the pharmacokinetic behavior of total platinum in biological matrices for other L-OHP delivery systems.

LC-DAD/ESI-MS/MS characterization of phenolic constituents in Rosa canina L. and its protective effect in cells.

In an aim to prove the efficiency of polyphenols of Rosa canina fruits in promoting human health. A methanolic extract of R. canina fruits was prepared by successive maceration with solvents of increasing polarity. The polyphenol composition was analyzed by HPLC-DAD-ESI-MS. The biological activity of this extract on SH-SY5Y cells and HepG2 cells was then studied. The antioxidant activity was tested by various in vitro tests such as DPPH-radical-scavenging activity, FRAP assay, hydroxyl radical scavenging assay and total antioxidant capacity. The subacute toxicity of R. canina was tested on female rats by repeated intraperitoneal administration of various doses. The phenolic profiles showed 25 antioxidants distributed into three classes of phenolic compounds: glycosylated and agglomerated flavonoids/isoflavonoids, tannins and phenanthrenes. Qualitative phytochemical analyses showed that this extract lacks alkaloids. The methanolic extract of R. canina fruits has a total antioxidant capacity of 82.69 ± 1.18 µg EAA/mg of methanol extract and the IC50 of the methods used is in the following increasing order: FRAP assay (61.88 µg/ml), then hydroxyl radical scavenging assay (67.45 µg/ml) and then DPPH radical-scavenging activity (129.81 µg/ml). The extract of R. canina did not cause any phenotypic signs of toxicity or mortality during and after treatment. The LD50 was >5,000 mg/kg, hence, R. canina was considered nontoxic. An in vivo study proved the protective effect of R. canina against cardiac and hepato-renal toxicities. These results drew the importance of a healthy diet, where diets rich in R. canina fruits can be used as a rich natural source of antioxidants and anticarcinogenic phenolic compounds.

Co-delivery of siRNA and hypericin into cancer cells by hyaluronic acid modified PLGA-PEI nanoparticles.

CONTEXT: Malignant tumors cause more death because of the resistance of the hypoxic cancer cell toward radiotherapy. Targeting for hypoxic cancer area and gene silencing to overcome the hypoxia are two kinds of important therapeutic strategies for treating tumors. OBJECTIVE: In order to explore the combined effects of gene therapy and hypericin (Hy) on tumor cells, hypoxia-inducible factor 1 alpha (HIF-1α) small interfering ribonucleic acid (siRNA) was transfected into the hypoxic human nasopharyngeal carcinoma (CNE2) cells using Hy-encapsulated nanocomplexes (Hy-HPP NPs) as a carrier which would achieve dual targeting to the tumor necrosis area. MATERIALS AND METHODS: NPs were prepared by emulsion-diffusion-evaporation method. Formulations were evaluated by conducting in vitro physicochemical studies, electrophoresis, in vivo study, and biochemical studies. RESULTS AND DISCUSSIONS: Hy-loaded nanoparticles with a mean size of around 160 nm was able to enhance the accumulation in the tumors by enhanced permeability and retention effect. The electrophoresis confirmed the good stability of siRNA/Hy-HPP NPs in the presence of phosphate-buffered saline (pH 7.4), competitive heparin, and RNase. The results of transfection showed that the uptake of siRNA was significantly increased up to 50% in CNE2 cells. The level of the HIF-1α with Hy-encapsulated nanocomplexes was significantly reduced to 30% in the transfected CNE2 cells. In vivo studies, the carrier exhibited higher intensity at the tumor tissue cells and higher affinity toward the necrotic tumor tissue. CONCLUSION: Results demonstrated that Hy-HPP NPs could significantly enhance the tranfection efficiency of siRNA, suggesting Hy-encapsulated nanoparticle as an efficient gene carrier. The co-delivery of HIF-1α siRNA (siHIF-1α) and Hy could efficiently decrease the level of HIF-1α and increase the affinity toward necrotic tissues. Hence, this is a promising strategy for further application in radiotherapy.

Antitumor activity of TNF-α after intratumoral injection using an in situ thermosensitive hydrogel.

Local drug delivery strategies based on nanoparticles, gels, polymeric films, rods and wafers are increasingly used in cancer chemotherapy in order to enhance therapeutic effect and reduce systemic toxicity. Herein, a biodegradable and biocompatible in situ thermosensitive hydrogel was designed and employed to deliver tumor necrosis factor-α (TNF-α) locally by intratumoral injection. The triblock copolymer was synthesized by ring-opening polymerization (ROP) of ß-butyrolactone (ß-BL) and lactide (LA) in bulk using polyethylene glycol (PEG) as an initiator and Sn(Oct)2 as the catalyst, the polymer was characterized by NMR, gel permeation chromatography and differential scanning calorimetry. Blood and tumor pharmacokinetics and in vivo antitumor activity of TNF-α after intratumoral administration in hydrogel or solution with the same dose were evaluated on S180 tumor-bearing mice. Compared with TNF-α solution, TNF-α hydrogel exhibited a longer T1/2 (4-fold) and higher AUCtumor (19-fold), but Cmax was lower (0.5-fold), which means that the hydrogel formulation improved the efficacy with a lower systhemic exposure than the solution formation. In addition, TNF-α hydrogel improved the antitumor activity and survival due to lower systemic exposure than the solution. These results demonstrate that the in situ thermosensitive hydrogel-based local delivery system by intratumoral injection is well suited for the administration of TNF-α.

Cationic polymer-based micro-emulgel with self-preserving ability for transdermal delivery of diclofenac sodium.

The objective of the present study was to develop a topical preparation with enhanced skin permeation, high safety and self-preserving ability. Microemulsion (ME) and cationic polymer based micro-emulgel (CPBM) were investigated for the transdermal delivery of diclofenac sodium (DS). Medium-chain triglyceride was selected as the oil phase of ME due to its good solubilization of DS and high safety. Orthogonal test was applied to optimize the formula of ME based on the cumulative skin permeation amount in vitro after preliminary formula test. Chitosan (CS) or polylysine was employed as the cationic polymer in the formula of CPBM. The transdermal delivery of DS was evaluated through in vitro skin permeation test. The results showed that the skin permeation rate of DS from the optimized CPBM (126.17 ± 15.82 µg/cm(2)/h) were 1.86-folds and 5.76-folds higher than that of DS commercial Emulgel and DS control hydrogel, respectively. MEs and the cationic polymer were found to have skin penetration co-enhancing effect when they were combined in the CPBM system. Furthermore, the CPBM showed a good growth inhibition of E. coli and S. aureus. The stability test revealed that the CPBM was stable at room temperature and 4 °C for a period of three months.

Novel designed polyoxyethylene nonionic surfactant with improved safety and efficiency for anticancer drug delivery.

In order to limit the adverse reactions caused by polysorbate 80 in Taxotere(®), a widely used formulation of docetaxel, a safe and effective nanocarrier for this drug has been developed based on micelles formed by a new class of well-defined polyoxyethylene sorbitol oleate (PSO) with sorbitol as the matrix in aqueous solution. The physicochemical properties of the amphiphilic surfactant and the resulting micelles can be easily fine-tuned by the homogeneous sorbitol matrix and pure oleic acid. Composition, critical micelle concentration, and entrapment efficiency were investigated by ultraviolet visible spectroscopy, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, fluorospectrophotometry, and high-performance liquid chromatography. In vitro and in vivo evaluation revealed that PSO had exceptionally low hemolysis and histamine release rates compared with commercial polysorbate 80. Moreover, the tumor targeting delivery of PSO was investigated by in vivo imaging in S180 tumor-bearing mice. The results suggest that this novel delivery system, PSO, provides an acceptable alternative to polysorbate 80 for delivery of docetaxel. Further, due to the hypoallergenic nature of PSO, the mechanism of pseudoallergy caused by the polyoxyethylene nonionic surfactant was investigated. Based on in vitro cell analysis, it was assumed that the initial contact of polyoxyethylene nonionic surfactant with mast cells provoked pseudoallergy via polyamine receptor-mediated endocytosis.

Novel pH-sensitive charge-reversal cell penetrating peptide conjugated PEG-PLA micelles for docetaxel delivery: in vitro study.

In order to create a pH-sensitive charge-reversal system for cell penetrating peptides (CPP) to prevent non-specific internalization of the drug; and concomitantly enhance the physical stability and tumor targetability of poly(ethylene glycol)-poly(d,l-lactide) (PEG-PLA) micelles, two sets of novel PEG-PLA micelles were developed. Cell penetrating decapeptide arginine-glycine (RG)5 and a pH-sensitive masking decapeptide histidine-glutamic acid (HE)5 were conjugated at the PEG free end to produce pH sensitive with peptides outside micelles (PHPO), while the pH sensitive with peptides inside micelles (PHPI) are the micelles obtained with the two peptides conjugated to the free end of the PLA block. The polymers were successfully synthesized and characterized by (1)H NMR and GPC. The mixed micelles were prepared and characterized for their loading efficiency, particle size and zeta potential. The surface charge of PHPO was greatly affected by the pH of the solution and (RG)5:(HE)5 ratio at the surface. The pH value of the solution at which the surface charge of PHPO reversed could be manipulated by the feed ratio of (RG)5-PEG-PLA (RGO) and (HE)5-PEG-PLA (HEO), hence, HEO:RGO molar ratio of 45:55 was selected for tumor targeting. Docetaxel (DTX) was sufficiently solubilized by DTX-PHPO with a loading efficiency of 90.18 ± 1.65%. At pH 7.4, DTX loaded mPEG-PLA (DTX-PM) (41.2 ± 0.3 nm), DTX-PHPO (195.3 ± 1.9 nm) and DTX-PHPI (190.9 ± 4.5 nm) showed sustained DTX release of less than 55% within 48 h. However, at pH 6.8 DTX-PHPI released 87.29 ± 0.24%, while DTX-PHPO released 70.49 ± 0.39% of the initial DTX amount within 48 h. Moreover, the physical stability of DTX-PHPO was increased due to the electrostatic interaction of the two peptides. The cellular uptake of DTX-PHPO in SGC-7901 cells and the cell killing effect tested on MCF-7 cells were enhanced by 2 folds at pH 6.8 compared to pH 7.4. Hence, DTX-PHPO is highly pH-sensitive in mildly acidic pH and exhibited higher internalization, but DTX-PHPI exhibited accelerated release. Meanwhile, both formulations displayed low internalization and release at pH greater than 7. This pH sensitive charge reversal design can offer a promising safe carrier using both CPPs and PEG-PLA micelles.

Synthesis, characterization, biodegradability and biocompatibility of a temperature-sensitive PBLA-PEG-PBLA hydrogel as protein delivery system with low critical gelation concentration.

Temperature-sensitive hydrogels were designed using a series of A-B-A triblock copolymers consisting of poly (ethylene glycol) (PEG) with different molecular weights as the hydrophilic block B and poly (ß-butyrolactone-co-lactic acid)(PBLA) with varying block lengths and composition as the hydrophobic block A. The triblock copolymers were synthesized by ring-opening polymerization (ROP) of ß-BL and LA in bulk using PEG as an initiator and Sn(Oct)2 as the catalyst. Their chemical structure and molecular characteristics were determined by NMR, GPC and DSC, and the relationship between structure and phase behaviors in aqueous solutions was investigated as well. It was found that the phase behaviors in aqueous solutions including critical micelle concentration (CMC), sol-gel-sedimentation phase transition temperature, gel window width and critical gelation concentration (CGC) are largely dependent on the molecular weight and block length ratio of PEG/PBLA. Most importantly, they show a very low CGC ranging from 4 to 8 wt% because of the introduction of ß-BL. Furthermore, the biodegradability and biocompatibility of the hydrogels were evaluated. Finally, lysozyme as a model protein was used to evaluate the ability to deliver protein drugs in a sustained release manner and biologically active form. All results demonstrated that the temperature-sensitive in situ forming hydrogel has a promising potential as sustained delivery system for protein drugs.

Development and characterization of stabilized double loaded mPEG-PDLLA micelles for simultaneous delivery of paclitaxel and docetaxel.

OBJECTIVE: Double loaded micelles (DLM) in which paclitaxel (PTX) and docetaxel (DTX) were co-solubilized with monomethoxy poly(ethylene glycol)-block-poly(d,l-lactide) (mPEG-PLA) copolymer were prepared and evaluated in an aim to investigate the effect of a combination of PTX and DTX on the stability of mPEG-PLA micelles compared to single drug-loaded micelles (SDM), especially that recent clinical anticancer formulations are limited by the existence of toxic excipients and stability issues. MATERIALS AND METHODS: The SDM and DLM of PTX and DTX were prepared by a solvent evaporation method. Micellar size, size distribution, drug loading content and drug release were investigated. Transmission electron microscopy was used to investigate the stabilization mechanism. RESULTS: The drug loading efficiency of both PTX and DTX in DLM and SDM were 25% and 10%, respectively. (1)H NMR showed a successful encapsulation of both drugs in the polymeric micelle. DLM showed better physical stability at drug concentrations higher than 1 mg/mL compared to SDM. Moreover, DLM, SDM-PTX and SDM-DTX were stable for 24, 9 and 1 h, respectively. The stabilization mechanism of DLM was investigated, a network structure of DLM was observed in TEM graphs. Furthermore, DLM showed complete and faster drug release compared to SDM. mPEG-PLA double loaded micelles can deliver two poorly water soluble anticancer drugs at clinically relevant doses. The obtained results offer a promising alternative for double drug therapy without any formulation associated undesirable effects and encourage further in vivo development and optimization of the DLM as a drug delivery system for anticancer drugs.

A poly(γ, L-glutamic acid)-citric acid based nanoconjugate for cisplatin delivery.

A cisplatin-loaded nanoconjugate, poly(γ, L-glutamic acid)-citric acid-cisplatin [γ-PGA-CA-CDDP], as a tumor-targeted drug delivery system with sustained release capacity was successfully synthesized and characterized, and its antitumor activity was evaluated. The particle size (107 ± 6.3 nm) and average molecular weight (66 kDa) were determined by dynamic light scattering (DLS) and gel permeation chromatography (GPC), respectively. The nanoconjugate delivery system released platinum in a sustained manner in PBS at 37 °C with an initial burst release during the first 8 h and 50% cumulative release within 48 h. Both in-vitro and in-vivo studies showed that the toxicity of γ-PGA-CA-CDDP nanoconjugate significantly decreased by comparison to that of unconjugated CDDP. The maximum tolerated dose (MTD) of γ-PGA-CA-CDDP nanoconjugate was about 38 mg/kg versus 8 mg/kg for CDDP. No apathy or acute adverse reactions were observed in γ-PGA-CA-CDDP nanoconjugate groups while mice expressed apathy at all dose levels with CDDP treatment. In ICR mice, the area under the curve and total body clearance values for γ-PGA-CA-CDDP nanoconjugate were 9-fold and one-twentieth of the values for CDDP, respectively. With the aid of near-infrared fluorescence (NIRF) imaging system, it was demonstrated that γ-PGA-CA-CDDP nanoconjugate gradually accumulated at the tumor site within 15 min postinjection and exhibited prolonged retention for more than 8 h. In H22-implanted mice, γ-PGA-CA-CDDP showed a significantly higher antitumor activity versus CDDP. These results reveal that γ-PGA-CA-CDDP nanoconjugate with improved stability, reduced toxicity and prolonged in-vivo retention time holds great potential in terms of clinical application to cancer therapy.

Novel oral delivery system of indomethacin by solidified mPEG-PDLLA micelles: in vivo study.

To administer indomethacin (IND) orally using polymeric micelles, IND loaded solidified polymeric micelles (IND-SPM) were prepared and evaluated for their in vitro and in vivo characteristics. IND and methoxy-poly(ethylene glycol) poly(d, l-lactide) copolymer (mPEG-PDLLA) were dissolved in acetone followed by the addition of an equivalent amount of polyplasdone XL-10 and stirred to obtain a suspension. Afterwards, acetone was completely evaporated. It was found that IND-SPM generates small polymeric micelles of 18.1 nm. Moreover, the solubility of IND at pH 6.8 increased 4.6-folds, and more than 90% of IND in 20 mg of IND-SPM was dissolved in 250 mL SIF pH 6.8 within 30 min. Pharmacokinetic parameters in fasted rats showed that IND-SPM 1:3 resulted in 3-folds increase of AUC and C(max) compared to commercial IND. mPEG-PDLLA micelles were found to be efficient carriers for oral administration of IND as solid dosage forms by adsorption on polyplasdone XL-10.

A novel tumor-targeted delivery system with hydrophobized hyaluronic acid-spermine conjugates (HHSCs) for efficient receptor-mediated siRNA delivery.

A novel target specific small interfering RNA (siRNA) delivery system was successfully developed using hydrophobized hyaluronic acid-spermine conjugates (HHSCs), which were previously synthesized and their properties were also characterized in our published papers. Fluorescein isothiocyanate (FITC) labeled specific Silencer Select siRNAs were used as a model system suppressing the cyclooxygenase-2 (COX-2) gene expression. The polymers were able to effectively bind siRNA, self-assemble into micelles, protect siRNA from degradation by nuclease and release complexed siRNA efficiently in the presence of low concentrations of polyanionic heparin. The cytotoxicity of siRNA/HHSCs complex to SGC-7901 cells was lower than that of siRNA/PEI 25k and Lipofectamine 2000 complex according to the MTT assay. When SGC-7901 and GES-1 cells were treated with FITC labeled siRNA/HHSCs complexes, SGC-7901 cells, with a cluster determinant 44 receptor (CD(44)), showed higher green fluorescent intensity than GES-1 cells because of the HA receptor mediated endocytosis of the complex. In addition, the inhibitory effect on the uptake in the presence of free HA in the transfection medium revealed that siRNA/HHSC-1 complex was selectively taken up to SGC-7901 cells via HA-receptor mediated endocytosis. Based on flow cytometry and microscopy, observation revealed that siRNA/HHSC complex was taken up preferentially through caveolae-mediated endocytosis, which may be a desirable pathway for avoiding the lysosomal degradation of delivered genes. All these results demonstrated that the intracellular delivery of siRNA/HHSC complex could be facilitated by the HA-receptor mediated endocytosis.

Solidified mPEG-PDLLA micelles as a novel oral delivery system of indomethacin / 药学学报

In this study, indomethacin (IND) loaded solidified-polymeric micelles (IND-SPM) were prepared. Their in vitro characteristics were investigated. Methoxy-poly(ethylene glycol) poly(D, L-lactide) copolymer (mPEG-PDLLA) was used as IND carrier. The preparation of IND-SPM was conducted by solution-absorption method and evaporation by rotary evaporator. Polyplasdone XL-10 was used as adsorbent. The solution-absorption method was conducted by the following procedure; IND and mPEG-PDLLA were dissolved in acetone, followed by addition of polyplasdone XL-10 and stirred to obtain a suspension. The powder of IND-SPM was simply obtained after the organic solvent was completely evaporated. More than 90% (w/w) of IND (20 mg) in the powder was dissolved in 250 mL PBS within 30 min. DSC, 1H NMR and SEM results proved that IND was encapsulated within mPEG-PDLLA. The solubility of IND in the system increased 4.6 times with the highest amount of copolymer. The solidified particles were found to be suitable for the formulation of tablets or capsules.

Solidified mPEG-PDLLA micelles as a novel oral delivery system of indomethacin / 药学学报

In this study, indomethacin (IND) loaded solidified-polymeric micelles (IND-SPM) were prepared. Their in vitro characteristics were investigated. Methoxy-poly(ethylene glycol) poly(D, L-lactide) copolymer (mPEG-PDLLA) was used as IND carrier. The preparation of IND-SPM was conducted by solution-absorption method and evaporation by rotary evaporator. Polyplasdone XL-10 was used as adsorbent. The solution-absorption method was conducted by the following procedure; IND and mPEG-PDLLA were dissolved in acetone, followed by addition of polyplasdone XL-10 and stirred to obtain a suspension. The powder of IND-SPM was simply obtained after the organic solvent was completely evaporated. More than 90% (w/w) of IND (20 mg) in the powder was dissolved in 250 mL PBS within 30 min. DSC, 1H NMR and SEM results proved that IND was encapsulated within mPEG-PDLLA. The solubility of IND in the system increased 4.6 times with the highest amount of copolymer. The solidified particles were found to be suitable for the formulation of tablets or capsules.