Precisely targeted drug delivery by mesenchymal stem cells-based biomimetic liposomes to cerebral ischemia-reperfusion injured hemisphere.

The precise and targeted delivery of therapeutic agents to the lesion sites remains a major challenge in treating brain diseases represented by ischemic stroke. Herein, we modified liposomes with mesenchymal stem cells (MSC) membrane to construct biomimetic liposomes, termed MSCsome. MSCsome (115.99 ± 4.03 nm) exhibited concentrated accumulation in the cerebral infarcted hemisphere of mice with cerebral ischemia-reperfusion injury, while showing uniform distribution in the two cerebral hemispheres of normal mice. Moreover, MSCsome exhibited high colocalization with damaged nerve cells in the infarcted hemisphere, highlighting its advantageous precise targeting capabilities over liposomes at both the tissue and cellular levels. Leveraging its superior targeting properties, MSCsome effectively delivered Dl-3-n-butylphthalide (NBP) to the injured hemisphere, making a single-dose (15 mg/kg) intravenous injection of NBP-encapsulated MSCsome facilitate the recovery of motor functions in model mice by improving the damaged microenvironment and suppressing neuroinflammation. This study underscores that the modification of the MSC membrane notably enhances the capacity of liposomes for precisely targeting the injured hemisphere, which is particularly crucial in treating cerebral ischemia-reperfusion injury.

Inorganic nanoparticle-integrated mesenchymal stem cells: A potential biological agent for multifaceted applications.

Mesenchymal stem cell (MSC)-based therapies are flourishing. MSCs could be used as potential therapeutic agents for regenerative medicine due to their own repair function. Meanwhile, the natural predisposition toward inflammation or injury sites makes them promising carriers for targeted drug delivery. Inorganic nanoparticles (INPs) are greatly favored for their unique properties and potential applications in biomedical fields. Current research has integrated INPs with MSCs to enhance their regenerative or antitumor functions. This model also allows the in vivo fate tracking of MSCs in multiple imaging modalities, as many INPs are also excellent contrast agents. Thus, INP-integrated MSCs would be a multifunctional biologic agent with great potential. In this review, the current roles performed by the integration of INPs with MSCs, including (i) enhancing their repair and regeneration capacity via the improvement of migration, survival, paracrine, or differentiation properties, (ii) empowering tumor-killing ability through agent loaded or hyperthermia, and (iii) conferring traceability are summarized. An introduction of INP-integrated MSCs for simultaneous treatment and tracking is also included. The promising applications of INP-integrated MSCs in future treatments are emphasized and the challenges to their clinical translation are discussed.

Cell membrane-coated nanoparticles: a novel multifunctional biomimetic drug delivery system.

Recently, nanoparticle-based drug delivery systems have been widely used for the treatment, prevention, and detection of diseases. Improving the targeted delivery ability of nanoparticles has emerged as a critical issue that must be addressed as soon as possible. The bionic cell membrane coating technology has become a novel concept for the design of nanoparticles. The diverse biological roles of cell membrane surface proteins endow nanoparticles with several functions, such as immune escape, long circulation time, and targeted delivery; therefore, these proteins are being extensively studied in the fields of drug delivery, detoxification, and cancer treatment. Furthermore, hybrid cell membrane-coated nanoparticles enhance the beneficial effects of monotypic cell membranes, resulting in multifunctional and efficient delivery carriers. This review focuses on the synthesis, development, and application of the cell membrane coating technology and discusses the function and mechanism of monotypic/hybrid cell membrane-modified nanoparticles in detail. Moreover, it summarizes the applications of cell membranes from different sources and discusses the challenges that may be faced during the clinical application of bionic carriers, including their production, mechanism, and quality control. We hope this review will attract more scholars toward bionic cell membrane carriers and provide certain ideas and directions for solving the existing problems.

The in vivo fate and targeting engineering of crossover vesicle-based gene delivery system.

Exosomes and biomimetic vesicles are widely used for gene delivery because of their excellent gene loading capacity and stability and their natural targeting delivery potential. These vesicles take advantages of both cell-based bioactive delivery system and synthetical lipid-derived nanovectors to form crossover characteristics. To further optimize the specific targeting properties of crossover vesicles, studies of their in vivo fate and various engineering approaches including nanobiotechnology are required. This review describes the preparation process of exosomes and biomimetic vesicles, and summarizes the mechanism of loading and delivery of nucleic acids or gene editing systems. We provide a comprehensive overview of the techniques employed for preparing the targeting crossover vesicles based on their cellular uptake and targeting mechanism. To delineate the future prospects of crossover vesicle gene delivery systems, various challenges and clinical applications of vesicles have also been discussed.

Secondary Packages cannot Protect Liquid Biopharmaceutical Formulations from Dropping-Induced Degradation.

PURPOSES: Liquid protein-based biopharmaceutical formulations have been reported to form aggregation and protein sub-visible particles (SbVPs) during dropping (Randolph et al., J Pharm Sci 2015, 104, 602). However, effects of secondary package on liquid biopharmaceutical formulation stability during dropping are overlooked and have not been reported so far. This study reports the first real-world evaluation on effects of secondary package on liquid biopharmaceutical formulation stability during dropping, using two monoclonal antibodies (mAb-1 and mAb-2) and one fusion protein (FP-1) as model biopharmaceuticals. METHODS: The potential protective effects of secondary package and formulation composition on liquid biopharmaceutical formulations during dropping were evaluated with micro-flow imaging (MFI) and dynamic light scattering (DLS). RESULTS: The dropping-induced degradation could be detected with the two sensitive particle analyzing techniques MFI and DLS. Formulation compositions have dramatic impact on biopharmaceutical stability during dropping. Surprisingly, unlike the primary packages that have been reported to impact liquid biopharmaceutical stability, the secondary packaging system as described in our current preliminary design has little or no protective effect during dropping. CONCLUSIONS: Our study is the first real-world data showing that the secondary package system has little to no effect on the liquid biopharmaceutical formulation quality during dropping. On the contrary, the stability of liquid biopharmaceutical formulations during dropping is more relevant to formulation compositions and primary packages.

Recent Progress in the Design and Medical Application of In Situ Self-Assembled Polypeptide Materials.

Inspired by molecular self-assembly, which is ubiquitous in natural environments and biological systems, self-assembled peptides have become a research hotspot in the biomedical field due to their inherent biocompatibility and biodegradability, properties that are afforded by the amide linkages forming the peptide backbone. This review summarizes the biological advantages, principles, and design strategies of self-assembled polypeptide systems. We then focus on the latest advances in in situ self-assembly of polypeptides in medical applications, such as oncotherapy, materials science, regenerative medicine, and drug delivery, and then briefly discuss their potential challenges in clinical treatment.

Active stealth and self-positioning biomimetic vehicles achieved effective antitumor therapy.

Mesenchymal stem cells (MSCs) are recognized as promising drug delivery vehicles. However, the limitation of drug loading capacity and safety considerations are two obstacles to the further application of MSCs. Here, we report MSC membrane-coated mesoporous silica nanoparticles (MSN@M) that maintain the active stealth and self-positioning drug delivery abilities of MSCs and resolve issues related to MSCs-mediated drug delivery. MSN@M was established through uniformly integrating MSC membrane onto a mesoporous silica nanoparticle (MSN) core by sonication. Reduced clearance of phagocytes mediated by CD47 marker on MSC membrane was observed in vitro, which explained the only ~ 25% clearance rate of MSN@M compared with MSN in vivo within 24 h. MSN@M also showed stronger tumor targeting and penetration ability compared with MSN in HepG2 tumor bearing mice. Simultaneously, MSN@M exhibited strong capacity for drug loading and sustained drug release ability of MSN when loaded with doxorubicin (DOX), the drug loading of MSN@M increased ~ 5 folds compared with MSC membrane. In HepG2 xenograft mice, DOX-loaded MSN@M effectively inhibited the growth of tumors and decreased the side effects of treatment by decreasing the exposure of other tissues to DOX. Consequently, our MSN@M may serve as alternative vehicles for MSCs and provide more options for antitumor treatment.

Comparative performance of FAS equation and Asian modified CKD-EPI in the determination of GFR in Chinese patients with CKD with the 99mTc-DTPA plasma clearance as the reference method.

BACKGROUND: Glomerular filtration rate (GFR) is a useful index in many clinical conditions. However, very few studies have assessed the performance of full age spectrum (FAS) equation and the Asian modified Chronic Kidney Disease-Epidemiology Collaboration (CKD-EPI) equation in the approximation of GFR in Chinese patients with chronic kidney disease. OBJECTIVE: This study aimed to compare the diagnostic performance of the above two creatinine-based equations. METHODS: A well designed single-center cross-sectional study was performed and the GFR was determined by 3 methods separately in the same day: technetium-99m-diethylene triamine pentaacetic acid (99mTc-DTPA) dual plasma sample clearance method (mGFR); FAS equation method; Asian modified CKD-EPI equation method. The gold standard method was the mGFR. Equations performance criteria considered correlation coefficient, bias, precision, accuracy and the ability to detect the mGFR less than 60ml/min/1.73m2. RESULTS: A total of 160 patients were enrolled. The diagnostic performance of FAS showed no significant difference in the correlation coefficient (0.89 vs 0.89), precision (15.9 vs 16.1ml/min/1.73m2), accuracy (75.0% vs 76.3%) and the ability to detect the mGFR less than 60ml/min/1.73m2 (0.94 vs 0.94) compared with the Asian modified CKD-EPI equation in all participants. The FAS showed a negative bias, while the new CKD-EPI equation showed a positive bias (-1.20 vs 1.30ml/min/1.73m2, P<0.001). However, they were all near to zero. In the mGFR<60ml/min/1.73m2 subgroup and mGFR>60ml/min/1.73m2 subgroup were consistent with that in the whole cohort. The precision and accuracy decreased when GFR>60ml/min/1.73m2 in both equations. CONCLUSIONS: The FAS equation and the Asian modified CKD-EPI equation had similar performance in determining the glomerular filtration rate in the Chinese patients with chronic kidney disease. Both the FAS equation and Asian modified CKD-EPI can be a satisfactory method and may be the most suitable creatinine-based equation.

Mitochondrial targeted doxorubicin derivatives delivered by ROS-responsive nanocarriers to breast tumor for overcoming of multidrug resistance.

Multidrug resistance (MDR) is a serious challenge in chemotherapy and also a major threat to breast cancer treatment. As an intracellular energy factory, mitochondria provide energy for drug efflux and are deeply involved in multidrug resistance. Mitochondrial targeted delivery of doxorubicin can overcome multidrug resistance by disrupting mitochondrial function. By incorporating a reactive oxygen species (ROS)-responsive hydrophobic group into the backbone structure of hyaluronic acid - a natural ligand for the highly expressed CD44 receptor on tumor surfaces, a novel ROS-responsive and CD44-targeting nano-carriers was constructed. In this study, mitochondria-targeted triphenylphosphine modified-doxorubicin (TPP-DOX) and amphipathic ROS-responsive hyaluronic acid derivatives (HA-PBPE) were synthesized and confirmed by 1H NMR. The nanocarriers TPP-DOX @ HA-PBPE was prepared in a regular shape and particle size of approximately 200 nm. Compared to free DOX, its antitumor activity in vitro and tumor passive targeting in vivo has been enhanced. The ROS-responsive TPP-DOX@HA-PBPE nanocarriers system provide a promising strategy for the reverse of MDR and efficient delivery of doxorubicin derivatives into drug-resistant cancer cells.

Protein Sub-Visible Particle and Free Radical formation of a Freeze-Dried Monoclonal Antibody Formulation During Dropping.

Dropping during shipping and handling of liquid biopharmaceutical formulations has long been known to cause protein degradation and aggregation. On the other hand, accidental dropping of freeze-dried protein formulations is generally considered not a major issue for biopharmaceutical quality. Reports of stability and especially the underling degradation mechanism(s) during shipping and handling of freeze-dried protein formulations were rarely seen in literature. In this manuscript, we report an interesting phenomenon in which repeated dropping of freeze-dried monoclonal antibody X (mAb-X) formulation powder resulted in significant protein sub-visible particles (SbVPs) in the reconstituted liquid as determined by the sensitive particle analyzing technique micro-flow imaging (MFI). Free radicals were observed after repeated dropping by electron paramagnetic resonance (EPR). Formation of SbVPs could be partially inhibited by the free radical scavengers methionine and 3-carbamoyl-2,2,5,5-tetramethyl-1-pyrrolidin-yloxy free radical (CTPO). The amount of free radicals and SbVPs was correlated to the sample temperature during dropping. Therefore we propose that the high temperature formed during dropping was probably the root cause for protein aggregation and free radical formation, which could further cause protein aggregation. Our observations suggest that similar to liquid protein formulations, dropping of freeze-dried protein formulations should also be avoided or mitigated.

Comparative performance of FAS equation and Asian modified CKD-EPI in the determination of GFR in Chinese patients with CKD with the 99mTc-DTPA plasma clearance as the reference method.

BACKGROUND: Glomerular filtration rate (GFR) is a useful index in many clinical conditions. However, very few studies have assessed the performance of full age spectrum (FAS) equation and the Asian modified Chronic Kidney Disease-Epidemiology Collaboration (CKD-EPI) equation in the approximation of GFR in Chinese patients with chronic kidney disease. OBJECTIVE: This study aimed to compare the diagnostic performance of the above two creatinine-based equations. METHODS: A well designed single-center cross-sectional study was performed and the GFR was determined by 3 methods separately in the same day: technetium-99m-diethylene triamine pentaacetic acid (99mTc-DTPA) dual plasma sample clearance method (mGFR); FAS equation method; Asian modified CKD-EPI equation method. The gold standard method was the mGFR. Equations performance criteria considered correlation coefficient, bias, precision, accuracy and the ability to detect the mGFR less than 60ml/min/1.73m2. RESULTS: A total of 160 patients were enrolled. The diagnostic performance of FAS showed no significant difference in the correlation coefficient (0.89 vs 0.89), precision (15.9 vs 16.1ml/min/1.73m2), accuracy (75.0% vs 76.3%) and the ability to detect the mGFR less than 60ml/min/1.73m2 (0.94 vs 0.94) compared with the Asian modified CKD-EPI equation in all participants. The FAS showed a negative bias, while the new CKD-EPI equation showed a positive bias (-1.20 vs 1.30ml/min/1.73m2, P<0.001). However, they were all near to zero. In the mGFR<60ml/min/1.73m2 subgroup and mGFR>60ml/min/1.73m2 subgroup were consistent with that in the whole cohort. The precision and accuracy decreased when GFR>60ml/min/1.73m2 in both equations. CONCLUSIONS: The FAS equation and the Asian modified CKD-EPI equation had similar performance in determining the glomerular filtration rate in the Chinese patients with chronic kidney disease. Both the FAS equation and Asian modified CKD-EPI can be a satisfactory method and may be the most suitable creatinine-based equation.

Silver Nanoparticles Induce DNA Hypomethylation through Proteasome-Mediated Degradation of DNA Methyltransferase 1.

Nanoparticles are used in many fields and in everyday products. Silver nanoparticles are the most frequently used nanoparticles; for example, in food-related products, owing to their antibacterial activity. However, it has been pointed out that they might have unexpected biological effects, and evaluation of their effects is underway. Although there is a growing body of evidence that nanoparticles can also induce epigenetic changes, there is still little information on the underlying mechanisms. Here, we evaluated changes in DNA methylation induced by silver nanoparticles and attempted to elucidate the induction mechanism. Immunofluorescence staining analysis revealed that silver nanoparticles with a diameter of 10, 50, or 100 nm (nAg10, nAg50, and nAg100, respectively) decreased the content of methylated DNA in A549 alveolar epithelial cells. The level of DNA methyltransferase 1 (Dnmt1) protein, which is involved in maintaining methylation during DNA replication, was significantly decreased, whereas that of Dnmt3b, which is responsible for de novo DNA methylation, was significantly increased by nAg10 treatment. Co-treatment with nAg10 and cycloheximide, which inhibits translation by inhibiting the translocation step of protein synthesis, decreased the level of Dnmt1 in comparison with nAg10-treated A549 cells, indicating a post-translational effect of nAg10. Furthermore, pretreatment with the proteasome inhibitor lactacystin restored the levels of Dnmt1 protein and DNA methylation in nAg10-treated cells. Collectively, these results suggest that nAg10 induced DNA hypomethylation through a proteasome-mediated degradation of Dnmt1.

Formation of an Unprecedented Impurity during CE-SDS Analysis of a Recombinant Protein.

PURPOSES: The main purposes of this article are to describe an unprecedented phenomenon in which significant amount of a shoulder peak impurity was observed during normal non-reducing capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) analysis of a recombinant fusion protein X, and to evaluate the root cause for this phenomenon. METHODS: A series of experiments were conducted to study the nature of this degradation. Effects of iodoacetamide (IAM), heating temperature, duration, and SDS on the formation of this specific impurity were evaluated using a variety of characterization techniques. RESULTS: The formation of the impurity as observed in CE-SDS was actually due to alkylation of lysine and serine residues with IAM, as confirmed by peptide mapping and LC-MS/MS, which increased the molecular weight and therefore decreased the electrophoretic mobility. The amount of impurity was also strongly dependent on sample preparation conditions including the presence or absence of SDS. CONCLUSIONS: Our study clearly suggested that even though IAM has been used extensively as an alkylation reagent in the traditional non-reducing CE-SDS analysis of monoclonal antibodies and other proteins, alkylation with IAM could potentially lead to additional impurity peak, and therefore complicating analysis. Therefore, before performing CE-SDS and other analyses, the effects of sample preparation procedures on analytical results must be evaluated. For protein X, IAM should be excluded for CE-SDS analysis.

Development and Evaluation of a System for the Semi-Quantitative Determination of the Physical Properties of Skin After Exposure to Silver Nanoparticles.

In order to ensure the safe usage of silver nanoparticles (nAgs) in cosmetics, it is necessary to reveal the physical properties of nAgs inside the skin, as these properties may change during the process of percutaneous absorption. In this study, we aimed to establish an analytical system based on single particle inductively coupled plasma mass spectrometry (sp-ICP-MS) to determine the physical properties of nAgs in the skin. First, we optimized a pretreatment method for solubilizing the skin samples and then showed that most of the nAgs were recovered by sodium hydroxide treatment while remaining in particle form. For separating the skin into the epidermis and dermis, we screened several conditions of microwave irradiation. The sp-ICP-MS analysis indicated that the application of 200 W for 30 s was optimal, as this condition ensured complete separation of skin layers without changing the physical properties of the majority of nAgs. Finally, we evaluated the in vivo application by analyzing the quantity as well as the physical properties of Ag in the epidermis, dermis, and peripheral blood of mice after exposing the skin to nAgs or Ag+. Subsequent sp-ICP-MS analysis indicated that nAgs could be absorbed and distributed into the deeper layers in the ionized form, whereas Ag+ was absorbed and distributed without a change in physical properties. This study indicates that in order to obtain a comprehensive understanding of the response of skin following exposure to nAgs, it is essential to consider the distribution and particle size of not only nAgs but also Ag+ released from nAgs into the skin.

Engineering bacterial outer membrane vesicles as transdermal nanoplatforms for photo-TRAIL-programmed therapy against melanoma.

Melanoma is an aggressive cancer with rapid progression, relapse, and metastasis. Systemic therapies for melanoma exhibit limited anticancer potential and high toxicity. Here, we developed the outer membrane vesicles derived from transgenic Escherichia coli, modified with αvß3 integrin peptide targeting ligand and indocyanine green (named as I-P-OMVs), to induce the transdermal photo-TRAIL-programmed treatment in skin melanoma.-OMVs, which are outer membrane vesicles derived from transgenic Escherichia coli, modified with αvß3 integrin targeting ligand and indocyanine green (named as I-P-OMVs), to induce the transdermal photo-TRAIL-programmed treatment in skin melanoma. I-P-OMVs exhibited excellent stratum corneum penetration and specificity to melanoma. Upon near-infrared irritation, I-P-OMVs not only induced photothermal-photodynamic responses against primary melanoma spheroids but also activated TRAIL-induced apoptosis in disseminated tumor cells, resulting in a complete eradication of melanoma. I-P-OMVs are the first nanoplatforms to induce transdermal photo-TRAIL-programmed therapy in melanoma with enhanced antitumor performance and high safety, having great potential in cancer therapy.

Current advances in stem cell-based therapies for hair regeneration.

Alopecia is resulted from various factors that can decrease the regeneration capability of hair follicles and affect hair cycles. This process can be devastating physically and psychologically. Nevertheless, the available treatment strategies are limited, and the therapeutic outcomes are not satisfactory. According to the possible pathogenesis of nonscarring alopecia, especially androgenetic alopecia, recovering or replenishing the signals responsible for hair follicle stem cells activation is a promising strategy for hair regeneration. Recently, stem cell-based therapies, especially those based on the stem cell-derived conditioned medium (CM), which is secreted by stem cells and is rich in paracrine factors, have been widely explored as the hair regenerative medicine. Several studies have focused on altering the composition and up-regulating the amount of secretome of the stem cells, thereby enhancing its therapeutic effects. Besides, stem cell-derived exosomes, which are present in the CM as message entities, are also promising for hair regrowth. In this review, the up-to-date progress of research efforts focused on stem cell-based therapies for hair regeneration will be discussed, including their therapeutic potentials with respective merits and demerits, as well as the possible mechanisms.

Formation of protein sub-visible particles during powder grinding of a monoclonal antibody.

We have observed an interesting phenomenon in which grinding of freeze-dried monoclonal antibody X (mAb-X) formulation powder resulted to significant protein sub-visible particles (SbVPs) in the reconstituted liquid, which could only be observed by sensitive particle analytical methods such as MFI and DLS. Effects of grinding temperature and the free radical scavengers methionine and 3-carbamoyl-2,2,5,5-tetramethyl-1-pyrrolidin-yloxy free radical (CTPO) on the formation of SbVPs were also evaluated. Free radicals were observed by EPR and the amount of free radicals was correlated to the sample temperature prior to grinding. Formation of SbVPs could be partially inhibited by methionine and CTPO. The amount of SbVPs formed was dependent on the amount of free radicals/sample temperature prior to grinding. At higher temperatures, more free radicals and SbVPs formed. Other than the previously known protein degradation due to high temperature formed during mechanical grinding, we propose an unreported and supplementary mechanism, i.e., the formation of free radicals (i.e., due to break of CO or CS bonds) in the dried state during mechanical grinding, leading to protein particle formation in the reconstituted solution. Our observation suggested that mechanical grinding of protein powder should be avoided or used cautiously (i.e., grinding temperature, strength and time) and the effects on radical and particle formation be fully evaluated.

AIE/FRET-based versatile PEG-Pep-TPE/DOX nanoparticles for cancer therapy and real-time drug release monitoring.

On account of the biological significance of self-assembling peptides in blocking the cellular mass exchange as well as impeding the formation for actin filaments resulting in program cell death, stimuli-responsive polypeptide nanoparticles have attracted more and more attention. In this work, we successfully fabricated doxorubicin-loaded polyethylene glycol-block-peptide (FFKY)-block-tetraphenylethylene (PEG-Pep-TPE/DOX) nanoparticles, where the aggregation-induced emission luminogens (AIEgen, TPE-CHO) can become a fluorescence resonance energy transfer (FRET) pair with the entrapped antitumor drug DOX to detect the release of drugs dynamically. This is the first successful attempt to detect and quantify the change of FRET signals in A549 cells via three methods to monitor the cellular uptake of nanoprobes and intracellular drug molecule release intuitively. As we proposed here, the combination of free DOX and the self-assembling peptide could achieve the synergistic anticancer efficacy. The multifunctional PEG-Pep-TPE/DOX nanoparticles may provide a new opportunity for combination cancer therapy and real-time detection of the drug release from stimuli-responsive nanomedicine.

Development and evaluation of a simultaneous and efficient quantification strategy for final prostanoid metabolites in urine.

Prostanoids (PNs) play critical roles in various physiological and pathological processes. Therefore, it is important to understand the alternation of PN expression profiles. However, a simultaneous and efficient quantification system for final PN metabolites in urine has not yet been established. Here, we developed and evaluated a novel method to quantify all final PN metabolites. By purification using a reverse phase solid phase extraction (SPE) column, the matrix effects against the final PGD2, PGE2, and PGF2α metabolites were low, and their accuracies were nearly 100%. The matrix effects against the final PGI2 and TXA2 metabolites were high using reverse phase SPE column purification alone. By applying a tandem SPE method that combined reverse phase and ion exchange SPE columns, the matrix effects decreased so that the accuracy was nearly 100%. To validate the reliability of the method, each final metabolite was quantified from mouse urine to which the PNs (PGD2, PGE2, and PGI2) were intravenously administered. As a result, the amounts of PN metabolites were correlated with those of the PNs administered to the blood in a dose-dependent manner. To validate the method using human samples, the urinary metabolites of Crohn's disease (CD, a PN-related disease) patients and healthy individuals were quantified. All five metabolites were successfully quantified. Only final PGE2 metabolite levels were significantly higher in CD patients than those in healthy individuals, so that the urinary metabolite profiles of CD patients is determined. In conclusion, we developed a novel method to quantify all final PN metabolites simultaneously and efficiently and demonstrated the practicality of the method using human CD patient samples.

Mechanism investigation of ethosomes transdermal permeation.

Ethosomes are widely used to promote transdermal permeation of both lipophilic and hydrophilic drugs, but the mechanism of interaction between the ethosomes and the skin remains unclear. In this work, it was exploded with several technologies and facilities. Firstly, physical techniques such as attenuated total reflectance fourier-transform infrared and laser confocal Raman were used and the results indicated that the phospholipids configuration of stratum corneum changes from steady state to unstable state with the treatment of ethosomes. Differential scanning calorimetry reflected the thermodynamics change in stratum corneum after treatment with ethosomes. The results revealed that the skin of Bama mini-pigs, which is similar to human skin, treated by ethosomes had a relatively low Tm and enthalpy. Scanning electron microscopy and transmission electron microscopy showed that the microstructure and ultrastructure of stratum corneum was not damaged by ethosomes treatment. Furthermore, confocal laser scanning microscopy revealed that lipid labeled ethosomes could penetrate the skin via stratum corneum mainly through intercellular route, while during the process of penetration, phospholipids were retained in the upper epidermis. Cell experiments confirmed that ethosomes were distributed mainly on the cell membrane. Further study showed that only the drug-loaded ethosomes increased the amount of permeated drug. The current study, for the first time, elucidated the mechanistic behavior of ethosomes in transdermal application from molecular configuration, thermodynamic properties, ultrastructure, fluorescent labeling and cellular study. It is anticipated that the approaches and results described in the present study will benefit for better design of drug-loaded ethosomes.