First, the role

of Sir2 as a key regulator of yeast lifespan was challenged.24 Second, at that time, no ortholog for PNC1 in other organisms was found. Therefore, the answer as to whether this model is unique only for yeast remains elusive. The first uncertainty about the role Sir2 plays in modulating replicative longevity in yeast via regulation of the rate of ERC formation arose from the observation that the lifespan extension by overexpressing Sir2 was strain-dependent.24 In addition, it was noticed that in the BY4742 yeast strain, mutation of fob1, which blocks the formation of ERCs, Inhibitors,research,lifescience,medical or Sir2 overexpression, together with DR has a cumulative effect on yeast lifespan.24 Put simply, DR extended the lifespan of fob1 or sir2 double mutation. Thus, at least in this yeast strain the effect of DR cannot be Sir2 or ERC-dependent, as an additional increase of lifespan was seen with each treatment. In response to these claims other groups have shown that in the absence of Sir2 another yeast sirtuin, Inhibitors,research,lifescience,medical Hst2, takes over and regulates the positive effect of DR on Inhibitors,research,lifescience,medical yeast lifespan via ERC formation.25 However, this was not the end of the debate. Soon after, two researchers published that in the BY4742 yeast strain, double mutation of sir2 and fob1 along with a mutation in one of the hst isoforms hst1/hst2/hst4 has no significant effect on the yeast lifespan.26

Treatment with DR extended the lifespan of these combinations. Notably, Inhibitors,research,lifescience,medical the role of Hst3 in this study was complex. Mutations in hst3 only, or triple mutation sir2hst3fob1,

have a small but significant effect on yeast lifespan. However, once combined with hst4 mutation, yeast lifespan was significantly reduced. Moreover, DR was not able to extend the lifespan of yeast carrying mutations in fob1 and all yeast sirtuins. Interestingly, the authors did not report whether DR can extend the lifespan Inhibitors,research,lifescience,medical in hst3hst4 double mutations. Thus, the role of hst3 in DR response remains elusive. Taken together, despite extensive research, the question whether Sir2 or other sirtuins regulate yeast lifespan TCL during DR via controlling ERCs formation is still under debate. Recent studies have reported that Sir2 regulates yeast replicative lifespan by additional rDNA-independent mechanisms. During cytokinesis, the majority of learn more proteins damaged due to oxidative stress are maintained in the mother cell. Nystrom and his associates showed that Sir2 is required for this asymmetric inheritance, and absence of Sir2 results in an inheritance of oxidatively damaged proteins and reduced capacity to respond to oxidative stress in daughter cells.27 Others have shown that an age-associated decrease in Sir2 protein levels is accompanied by an increase in histone H4 K16 acetylation and regions.

Different strategies have been exploited with interesting results, for example, in the preparation of bioconjugates obtained by covalently linking HA to a cytotoxic drug such as, for example paclitaxel [39, 40] or doxorubicin [41, 42]. These topics are out of the scope of this paper where only strategies consisting in the design of HA learn more decorated nanosystems will be discussed in depth. 3. Chemical Conjugation of HA to Lipid-Based Nanocarriers Different approaches can be used to bind HA to the lipid-based nanocarriers, depending on the molecular weight of the HA as well as on the need to start from preformed nanocarriers

or from Inhibitors,research,lifescience,medical pure lipids that will be then used to prepare particles. HA binding to preformed nanocarriers was the firstly used method [43] and offers the advantage to conjugate the HA only on the external surface of the particle. Inhibitors,research,lifescience,medical Of course, this approach makes difficult the control of the density of attachment of HA on the carrier surface. Moreover, the lower specificity of the linkage, due to the possibility to bind different amino groups, results in a consequent multipoint attachment of the polymer on the nanocarrier that is then difficult to characterize. Alternatively, HA can be previously conjugated

to a pure lipid and then added in the lipid mixture during the preparation of the nanoparticles. Inhibitors,research,lifescience,medical This procedure permits the introduction of a controlled amount of HA on nanocarriers, but could require a more elaborated synthetic method. 3.1. HA Binding to Preformed Nanocarrier Inhibitors,research,lifescience,medical High molecular weight (HMW) HA was attached to the surface of preformed liposomes through amidation reaction between the aminoreactive group of a lipid on the

liposome surface, generally a phosphatidylethanolamine (PE), and HA glucuronic carboxylate (Figure 2) [13, 14, 43]. The amidation reaction was performed preactivating HA by incubation with the Inhibitors,research,lifescience,medical 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) condensing agent in acidic medium and then adding the activated HA to the nanocarrier suspension in a basic Endonuclease medium. Elimination of the excess of reagent and reaction byproducts was obtained by centrifugation and repeated washing. Figure 2 Strategies to prepare HA-coated nanocarriers. A schematic representation. (a) HA binding to preformed nanocarrier. Amidation reaction between HA-carboxyl group and aminoreactive group of lipid on the liposome surface. (b) Synthesis of HA-PE conjugates … 3.2. Preparation of HA-PE Preformed Conjugates HA conjugation to the lipid before nanocarrier preparation was carried out with both high and low molecular weight (LMW) polymers [12, 19]. In all cases, HA reacted with an aminoreactive group present on the lipid that was PE, also in this case (Figure 2). Two different conjugation methods have been proposed depending on the HA molecular weight.

This means to accommodate both their vulnerability and their abilities to act, and thereby make them feel valued as respected agents and human beings in the transition process. Authors’ contributions First author designed the study. Both authors collected the data.

First author performed the structural PERK inhibitor analysis and drafted the manuscript. Second author contributed to the interpretation of the findings and critical review of the manuscript. Both authors read and approved the final manuscript. Acknowledgements The authors thank the next of kin who participated in this study and the leaders in the home care services in two municipalities in Telemark for their kind help to recruit the participants. Conflict of interest and funding The authors report no conflict

of interest. This work was funded by Telemark University College. Note 1.In Norway most home care services are organized in line with the purchaser–provider model, which is inspired by the New Public Management movement. The purchaser unit is a separate unit in the municipality health care services, where the responsibility for needs assessment is placed. The responsibility for exercising the care and re-examining the care VE 821 needs is however placed in home care (Vabø, 2012).
Phenomenography is a qualitative research approach which was developed in the 1970s within an educational framework, focusing on the qualitative experience of learning. Originally developed in Sweden, phenomenography studies the variations in ways that people look upon, experience, or understand phenomena in the world around them (Marton & Booth, 1997). It rests on

a non-dualistic ontological perspective, that the real objective world and a subjective world of mental representations are not separate (Marton, 2000). Instead, the world is “simultaneously objective and subjective,” which means that an experience “is as much an aspect of the object as it is of the subject” (Marton, 2000, p. 105). In the same manner, a phenomenon, or object of experience, is seen as a complex of the different ways of in which it can be experienced. These different ways are related to each other because they are experiences of the same object (Marton, 2000). Phenomenography is not to be mistaken for phenomenology, a well-established qualitative method used within healthcare research for many years, which investigates phenomena in order to reveal their structure or logic, and essence (Giorgi, 1999; Sjostrom & Dahlgren, 2002). In a phenomenographic analysis of interview texts, ways of experiencing a phenomenon are identified and classified under categories of description which often form a hierarchy, and which eventually can be defined in terms of increasing complexity (Marton & Booth, 1997).

The two groups have no significant difference in sex (chi-square test, P > 0.05), age analyses of variance (ANOVA, P > 0.05), and education (two samples t-test, P > 0.05). The detailed demographics of these two groups are shown in Table ​Table11. Table 1 Subject demographics All patients met the following inclusion criteria: (1) Current MDD attack as assessed by two experienced psychiatrists using Inhibitors,research,lifescience,medical the Structural Clinical this website Interview for Diagnostic and Statistical Manual of Mental Disorders (DSM-IV); (2) 18–45 years of age; (3) right-handed Han Chinese; (4) HAMD scores of at least 17. Patients and healthy controls were excluded if they had any of the following: (1) a history of neurological diseases or other serious physical diseases;

(2) a history of electroconvulsive therapy; (3) history of substance abuse (i.e., drugs, alcohol, and other psychoactive substance); (4) any contraindications for MRI. This study was approved by the Ethics Committee of the Second Xiangya Hospital, Central South University, Hunan, China. Written informed consent was obtained

Inhibitors,research,lifescience,medical from all subjects. Imaging acquisitions and data preprocessing All image data were acquired using a 1.5T Siemens GE Signa Twinspeed Scanner (General Electric Medical System, Milwaukee, WI). The resting-state Inhibitors,research,lifescience,medical fMRI was performed with a gradient-echo echo planar sequence. Subjects were asked to relax and think of nothing in particular with eyes closed but were requested not to fall asleep. A total of 180 volumes of echo-planar imagings were obtained axially (repetition time, 2000 msec; echo time, 40 msec; slices, 20; thickness, 5 mm; gap, 1 mm; field of view (FOV), 24′24 cm2; resolution, 64′64; Inhibitors,research,lifescience,medical flip angle, 90 degrees). Prior to preprocessing, the first 10 volumes were discarded to allow for scanner stabilization and the subjects’ adaptation to the environment. fMRI data preprocessing was then conducted by SPM8 (http://www.fil.ion.ucl.ac.uk/spm) and a data processing assistant for resting-state fMRI (DPARSF) (Yan and Zang 2010).

Briefly, the remaining functional scans were first Inhibitors,research,lifescience,medical corrected for within-scan acquisition time differences between slices, and then realigned to the middle volume to correct for interscan head motions. Subsequently, the functional scans were spatially normalized PDK4 by using T1 image unified segmentation (done by DPARSF directly) and resampled to 3 × 3×3 mm3. After normalization, the BOLD signal of each voxel was first detrended to abandon a linear trend and then passed through a band-pass filter (0.01–0.08 Hz) to reduce low-frequency drift and high-frequency physiological noise. Finally, nuisance covariates including head motion parameters, global mean signals, white matter signals, and cerebrospinal signals were regressed out from the BOLD signals. The masks of three ROIs related to the hate circuit (left SFGdor, right INS, and right PUT) were generated using the free software WFU_PickAtlas (http://www.ansir.wfubmc.edu) (Maldjian et al. 2003).

Indeed other neurotransmitters or neuromodulators have been

linked to depression. For example, therapeutic effects have been found by increasing dopamine levels using agomelatine (melatonergic MT1 and MT2 agonist and 5HT2C antagonist) [Den Boer et al. 2005]. Substance P (SP: neurokinin; NK) has also been linked to depression, which is logical seeing that NK1 receptors Inhibitors,research,lifescience,medical are colocalized with 5HT neurons in the dorsal raphe. Electrophysiological studies in mice stirred excitement as SP NK1 receptor antagonists, such as MK-869, desensitised 5HT autoreceptors within 3 hours. It was thought that this may remove the problem of delayed therapeutic onset [Blier et al. 2004], however the problem of delayed therapeutic onset remained perhaps attributable to species differences [Kramer et al. 1998]. Nonetheless, it is clear that factors beyond monoamines contribute to depression, highlighting a clear reason for the limited efficacy of antidepressants. The future of depression treatment Pharmacogenetics, how and why an individual Inhibitors,research,lifescience,medical responds to a Inhibitors,research,lifescience,medical given compound, is a nascent field in pharmacology generally and has offered potential to predict and optimize individual responses to medication. An example in the pharmacotherapy depression was identification in the STAR*D project of polymorphisms associated with favourable antidepressant

response, e.g. a functional polymorphism: 5HTTLPR, of the 5HT transporter gene (5HTT) has a short and long allele. The long allele has been associated with

better response to SSRIs, whereas the short allele with poorer response in Whites [Schosser and Inhibitors,research,lifescience,medical Kasper, 2009]. This fits in with genetic research from Caspi and colleagues that individuals heterozygous or homozygous for the short allele are more susceptible to depression following stressful events [Caspi et al. 2003]. Future work in the pharmacogenetics field has potential for changing depression management through the use of biomarkers and Inhibitors,research,lifescience,medical genotypes, which may permit early identification of whether an individual will gain sufficient benefits from antidepressants to justify the risks they entail. However, this is far from being achieved and significant problems are inherent in this field, e.g. differences in ethnicity as in the Asian population the short allele of 5HTTLPR polymorphism is associated with Dichloromethane dehalogenase better response [Kim et al. 2000]. Circumspection is required at this time, and excitement around early findings has been tempered by a failure to replicate these findings [Leuchter et al. 2009]. This individualized approach may help overcome the problem of heterogeneity, as currently the disorder of depression is too broad to serve as a useful construct for treatment development. Depression may not be a unitary disorder, but AUY-922 nmr rather an umbrella construct harvesting multiple disorders with varying biological pathophysiology which each require different treatment.

Table 2 CTT2-SL liposomes were made by pipetting the above-mentioned lipid mixture except the CTT2-PEG-lipid, to a round bottomed flask, dried under nitrogen and lyophilized for 2h to remove trace amounts of chloroform. Doxorubicin liposomes were prepared by using standard pH Screening Library research buy gradient technique [1]. To synthesize CTT2-PEG-3400-DSPE Caelyx/doxil-liposomes, CTT2-PEG-DSPE (1mg) was suspended in 400μl of buffer (100mM histidine, 55mM sucrose, pH 6.5), and 100μl of this CTT2-PEG-DSPE micelle suspension was added to 1ml Doxil/Caelyx solution or internally prepared similar to doxil-liposomes (Ortho Biotech).

In vivo murine studies were performed after incubating the mixture Inhibitors,research,lifescience,medical for 30min at 60°C. The incorporation efficiency, the percentage of total activity contained

in the liposome fractions, was measured by using radioisotope-labeled peptide and gel filtration to separate the unreacted micelle from the liposome; optimal reaction conditions Inhibitors,research,lifescience,medical were found to be 60°C at 30min (nearly 100% efficient). The doxorubicin leakage from the liposomes after the incorporation experiments was determined by comparing the amount of free doxorubicin versus liposome-bound doxorubicin before and after the experiment. The leakage was found to be minimal (the leakage before the incorporation Inhibitors,research,lifescience,medical was in average 4.5% and after Inhibitors,research,lifescience,medical the reaction in average 4.2%). 2.4. Radiolabeling of Peptides Radiolabeling of peptides and all liposomal formulations with iodine-125 (125I) was performed using the IODOGEN (Pierce, Rockford, IL). The CTT2-PEG3400-DSPE peptide was labeled with 125I using iodogen as a catalyst. 5MBq of Na125I (Amersham, Buckinghamshire, England) in 0.5ml PBS was added to a tube containing Inhibitors,research,lifescience,medical 10μg dried iodogen and 100μg CTT2-PEG3400-DSPE peptide construct. The mixture was incubated for 20min at room temperature. The 125I-bound

particle fractions were purified by elution from PD-10 columns. The activity of the peptide was determined in a gamma counter (Cobra II, Packard Instruments). 2.5. Animal Models and Tumor Inoculation The mice were cared for according to the instructions of the animal facility, and the experiments were approved next by an ethical committee of Helsinki University, Finland. Male athymic nu/nu mice (6–8weeks old, Harland) were provided with water and maintained on regular diets ad libitum. Subcutaneous human serous ovarian carcinoma (OV-90) xenograft models were generated by coinjecting equal volumes of cells (~5×106/100μl phosphate buffered saline, PBS) and matrigel subcutaneously into the hindlegs of nude mice. Average tumor volumes of 65mm3–200mm3were used for all studies. 2.6. In Vivo Biodistribution and Pharmacokinetics Following single i.v.

Sustained VT or even VF can follow the iatrogenic VT induced by rapid ventricular pacing, particularly in patients with

preoperatively compromised left ventricular function. Of course, VT or VF can always be indicative of severe coronary ischemia during the intervention. Patients who have received an implantable cardioverter defibrillator prior to TAVI should have the antitachycardia algorithms turned off during Inhibitors,research,lifescience,medical the intervention so as not to interfere with the episodes of rapid ventricular pacing. Conclusion While TAVI is a promising therapy for high-risk patients who are not candidates for traditional open surgery, the procedure has inherent challenges that must be overcome before it can be considered a truly safe alternative. It is the responsibility of the heart team to collectively work towards decreasing the complication rate of TAVI and ensuring a safe and effective alternative therapy for patients. Conflict of Interest Disclosure: All authors have completed and submitted Inhibitors,research,lifescience,medical the Methodist DeBakey Cardiovascular Journal Conflict of Interest Statement and the following was reported: Dr. Laborde Inhibitors,research,lifescience,medical is a consultant for Medtronic, Inc. Funding/Support: The authors have no funding disclosures. Contributor Information Jean-Claude Laborde, St. George’s Hospital, London, United Kingdom. Stephen J.D. Navitoclax in vivo Brecker, St. George’s Hospital, London, United Kingdom. David Roy, St.

George’s Hospital, London, United Kingdom. Marjan Jahangiri, St. George’s Hospital, London, United Kingdom.

Introduction Cardiovascular diseases (CVDs) claim more lives each year than cancer, chronic lower respiratory disease, and accidents combined. Clearly, there is a need for new therapies to treat Inhibitors,research,lifescience,medical this pervasive problem. The use of stem cell therapy in CVDs for protection, restoration, and regeneration has gathered momentum in the past few years.1–5 A variety of cell types have been considered as candidates.6 Currently available routes for delivering progenitor cells to the heart, which include intravenous (IV), intracoronary (IC), or direct epicardial injection and, more recently, injection in the coronary sinus, are Inhibitors,research,lifescience,medical inefficient due to low cell retention and a lack of targeted localization. Although IV delivery

of cells is the least invasive of these methods, most of the delivered cells are trapped in the lungs, with less than 1% homing to the infarcted Electron transport chain heart. During angioplasty, cells can be delivered by IC infusion directly to the region of interest. However, studies show that 50% to 90% of injected cells are lost by extrusion and that 90% of the remaining cells die within 1 week of implantation. Upon restoration of blood flow, the majority of cells are washed away from the region of interest, and only 3% of the delivered cells engraft into the heart. By comparison, some studies showed that direct intramuscular injection of cells into the heart wall resulted in a modest increase in the number of cells delivered to the myocardium, with 11% of the cells engrafting.

Nevertheless, early theories

of depression discounted the validity of the disorder in youth, suggesting that the necessary psychological mechanisms were not yet present for the experience of depression, or that depression was “masked” in the form of other disorders. In particular, it was considered that children did not have a well-developed superego. In 1975, the National Institute of Mental Health convened a meeting to discuss the incidence and diagnosis of depression in children. When the Inhibitors,research,lifescience,medical existence of depression in children became acceptable and the basic diagnostic criteria were established,1 research on Kinase Inhibitor Library concentration childhood depression burgeoned, resulting in the growth of theoretical models as well as empirical databases, and depression is no Inhibitors,research,lifescience,medical longer considered “an adult disease.” Despite this burgeoning research, some obstacles remained with regard to the pursuit of knowledge on adolescent depression. The early “storm and stress” theories of development suggested that many of the problems associated with depression, such as sadness, irritable mood, self-doubt, and social withdrawal, were normative expressions of adolescent

angst.2 It is now established, however, that many youngsters do not go through such emotional distress, and that adolescent depression is a serious disorder, often heralding chronic or recurrent problems into adulthood. A developmental framework in understanding childhood and adolescent depression Inhibitors,research,lifescience,medical In the past three decades, depression research in children and adolescents Inhibitors,research,lifescience,medical has progressed from applying simple extensions of clinical descriptions and theories developed in adults to generating an increasingly sophisticated understanding

of these disorders informed by the emerging field of developmental psychopathology. Research adopting this framework has taken into account the normative developmental processes influencing differences in the etiology, phenomenology, correlates, and outcomes of depression Inhibitors,research,lifescience,medical in children, adolescents, and adults.3-7 It is important to note, however, that this new field of research often does not differentiate among particular Sodium butyrate stages of development through childhood and adolescence. Although some continuity is likely across childhood and adolescence in the experience and expression of depression, the underlying risk mechanisms and the consequences of depression, some differences are also plausible. When applying a developmental perspective to psychopathology, one important issue to consider is the conceptualization of different life stages. For example, the transition from childhood to adolescence involves changes in multiple domains, including physical, sexual, cognitive, and social development, with a considerable range of individual differences in the age at which each of these changes occur. At present, there is no consensus on the clear boundaries in defining child and adolescent populations.

Only a few studies in postmortem brain tissue on a relatively small number of subjects have attempted to estimate the number of neurons in such subcortical structures as hypothalamus,

dorsal raphe nucleus, locus ceruleus, and amygdala.44-52 Results of these subcortical histopathological studies are somewhat inconsistent. Increases, decreases, or no change in the cell number or density are reported in the hypothalamus and brain stem nuclei in depressed subjects. Stereological investigation of specific types of hypothalamic neurons reveals an increase in the numbers of arginine-vasopressin (AVP)-immunoreactive neurons, oxytocin-expressing neurons, Inhibitors,research,lifescience,medical and corticotropin-releasing hormone (CRH) neurons in the paraventricular nucleus in subjects with BPD or MDD, compared to normal controls.44,45 Moreover, Inhibitors,research,lifescience,medical increases in CRH mRNA, and in the number of CRH neurons colocalizing AVP are also found in depressed patients.46,47 These findings

of increases in specific immunoreactive neurons arc consistent with the evidence of activation of the hypothalamicpituitary-adrenal (HPA) Inhibitors,research,lifescience,medical axis in some subsets of depressed patients.48 On the other hand, decreased number and density of nitric oxide synthase-containing neurons in the paraventricular hypothalamic nucleus are described in a small group of subjects with either MDD or BPD.49 Subtle structural abnormalities have been reported in mood disorders in the monoaminergic brain stem nuclei, the major sources of serotonin (dorsal raphe nucleus) and norepinephrine (locus Inhibitors,research,lifescience,medical ceruleus) projections to the cerebral cortex. An increased number and density

of tryptophan hydroxylase immunoreactive neurons is observed in the dorsal raphe nucleus of suicide victims with MDD compared with controls.50 In suicide victims, Arango et al51 report fewer pigmented neurons within the rostral locus ceruleus. Another study in a larger number of subjects found no differences in the number Inhibitors,research,lifescience,medical of isothipendyl pigmented neurons in the locus ceruleus between subjects with MDD (most were suicides) and control subjects.52 Although the number of neurons in the locus ceruleus does not appear altered in MDD, CRH immunoreactivity is increased in the locus ceruleus and pontine dorsal and median raphe nuclei.53,54 No VX-770 concentration changes in neuronal densities were detected in amygdala in subjects with either M’DD or BPD, as compared to normal controls.17 These postmortem findings suggest that some changes in the morphology of hypothalamic neurons and brain stem neurons may take place in mood disorders. However, future studies employing stereological techniques and a larger number of subjects are required to determine the exact pathology in these regions in depression.

As a result, also patients will benefit from these investments, in terms of innovative techniques, therapies, devices, and drugs designed to extend and improve their lives.
Drug delivery systems (DDSs) are useful for reducing drug side effects and maximizing drug action. The design of drug Ku0059436 carriers for DDS is the most important activity in this area. The advent of molecular biology studies has enabled the identification of many disease-causing proteins. Because some of these are effective as drugs, protein delivery systems Inhibitors,research,lifescience,medical have become important in DDS. A variety of nanoparticles such as liposomes, micelles, and polymers have been adopted as drug carriers [1–4]. Because polymers are

similar in size to proteins, they are not suitable as protein carriers. Whereas liposomes and micelles are larger than polymers and proteins, they can be used as protein carriers. However, because these are self-assembled nanoparticles, some treatments are Inhibitors,research,lifescience,medical necessary for their preparation as carriers. Since most proteins are sensitive to temperature, pH, and organic solvents, it is possible that such treatments induce protein denaturation. Hydroxyapatite

(HA), Ca10(PO4)6(OH)2, is a major component of hard tissues such as bones and teeth Inhibitors,research,lifescience,medical and has been used as a biomaterial [5, 6]. Because it has been reported that some proteins, such as bovine serum albumin (BSA) and lysozyme, can bind to HA just by mixing, it is a good candidate for a protein carrier [7–9]. In this study, we investigated the association and dissociation behavior of two bioactive proteins, cytochrome c and insulin, to HA. It is known that the release of cytochrome c from mitochondria

to the cytosol induces apoptosis. Therefore, the delivery of cytochrome c into the cytosol Inhibitors,research,lifescience,medical of cancer cells should Inhibitors,research,lifescience,medical induce apoptosis, which may be useful for cancer therapy [10]. Insulin, a key protein of diabetes, is commonly injected into diabetic patients to suppress blood sugar levels [11], and its controlled release can markedly improve their quality of life. Because the delivery of these two proteins is important, we attempted to use HA for a delivery system. The absorption and desorption on HA were affected by the surface conditions dependent on the preparation procedure of HA. Considering universal use of HA for protein delivery, commercially available HA was used as a carrier in this study. 3-mercaptopyruvate sulfurtransferase 2. Materials and Methods HA nanoparticles were purchased from Sigma-Aldrich (MO, USA). According to the material data sheet (no. 677418), the size and surface area were smaller than 200nm and larger than 9.4m2/g, respectively. Cytochrome c and insulin were obtained from Nacalai Tesque Inc. (Kyoto, Japan). Physicochemical properties of proteins used in this study are listed in Table 1. Cytochrome c is cationic, and insulin is anionic at physiological pH, and the molecular weight of cytochrome c is larger than that of insulin. Table 1 Proteins used in this study.