Synthetic molecular evolution of antimicrobial peptides.

As we learn more about how peptide structure and activity are related, we anticipate that antimicrobial peptides will be engineered to have strong potency and distinct functions and that synthetic peptides will have new biomedical applications, such as treatments for emerging infectious diseases. As a result of the enormous number of possible amino acid sequences and the low-throughput nature of antimicrobial peptide assays, computational tools for peptide design and optimization are needed for direct experimentation toward obtaining functional sequences. Recent developments in computational tools have improved peptide design, saving labor, reagents, costs, and time. At the same time, improvements in peptide synthesis and experimental platforms continue to reduce the cost and increase the throughput of peptide-drug screening. In this review, we discuss the current methods of peptide design and engineering, including in silico methods and peptide synthesis and screening, and highlight areas of potential improvement.

Predicting Membrane-Active Peptide Dynamics in Fluidic Lipid Membranes.

Understanding the interactions between peptides and lipid membranes could not only accelerate the development of antimicrobial peptides as treatments for infections but also be applied to finding targeted therapies for cancer and other diseases. However, designing biophysical experiments to study molecular interactions between flexible peptides and fluidic lipid membranes has been an ongoing challenge. Recently, with hardware advances, algorithm improvements, and more accurate parameterizations (i.e., force fields), all-atom molecular dynamics (MD) simulations have been used as a "computational microscope" to investigate the molecular interactions and mechanisms of membrane-active peptides in cell membranes (Chen et al., Curr Opin Struct Biol 61:160-166, 2020; Ulmschneider and Ulmschneider, Acc Chem Res 51(5):1106-1116, 2018; Dror et al., Annu Rev Biophys 41:429-452, 2012). In this chapter, we describe how to utilize MD simulations to predict and study peptide dynamics and how to validate the simulations by circular dichroism, intrinsic fluorescent probe, membrane leakage assay, electrical impedance, and isothermal titration calorimetry. Experimentally validated MD simulations open a new route towards peptide design starting from sequence and structure and leading to desirable functions.

Autism spectrum disorder: An examination of sex differences in neuropsychological and self-report measures of executive and non-executive cognitive function.

LAY ABSTRACT: Research comparing females and males with a diagnosis of autism suggests that there are sex differences in some characteristics such as behaviour regulation. One area not studied in detail is whether females and males with autism perform differently in tests of cognitive ability. The results of previous research are quite mixed. One explanation may be that some research comparing females and males with autism did not include a neurotypical control group for comparison. As a result, it is not clear whether the sex differences in cognitive ability observed in people with autism are similar to differences between neurotypical males and females. To better understand whether there are unique differences between males and females with autism, it is important to also compare them with neurotypical males and females. In our research, we included a neurotypical group and compared males and females with and without a diagnosis of autism. We found that the sex differences in autism are similar to what we observe in males and females without autism. Our study showed that compared with males, females (with and without autism) do better in assessments of processing speed, cognitive flexibility, verbal learning and memory and semantic fluency. Our results suggest that although females show different cognitive performance to males, these sex differences were not specific to the group with a diagnosis of autism.

Distress, quality of life and disability in treatment-seeking young adults with social anxiety disorder.

AIM: This study aimed to: (a) examine whether treatment-seeking young adults with social anxiety disorder (SAD) demonstrate similar degrees of distress, quality of life (QoL) and disability to those with other mental disorders; and (b) investigate the impact of comorbidity, specific comorbid conditions and antidepressants use on distress, QoL and disability in treatment-seeking young adults with SAD. METHODS: A cohort of treatment-seeking young adults (aged 16-45) diagnosed with SAD (N = 298) or other mental health disorders (N = 842; including depression, N = 349; bipolar, N = 141; psychosis, N = 173) completed self-report assessments of distress, QoL and disability. RESULTS: Young adults with SAD showed distress and disability of similar degree to those with most other mental disorders. Specifically, young adults with SAD reported significantly lower QoL than those with major depressive disorder or obsessive-compulsive disorder. Furthermore, young adults with SAD had the most difficulties in getting along with others and the second highest level of distress in comparison to other psychiatric groups. In comparison to antidepressants use, the presence of comorbidity showed a substantial negative influence on these health outcomes, particularly when presenting with comorbid depression or obsessive-compulsive disorder. CONCLUSIONS: Findings highlight significant impairments in young adults seeking treatment for SAD and the important moderating influence of comorbidity. This emphasizes the urgent need for effective management and treatment for its presentation and comorbidities in mental health services targeting young adults.

Validation of the 21-item Depression, Anxiety, and Stress Scales (DASS-21) in individuals with autism spectrum disorder.

The purpose of the study was to examine the internal consistency and validity of the 21-item Depression Anxiety Stress Scale (DASS-21) in individuals with Autism Spectrum Disorder (ASD) and without intellectual disability (IQ >= 70). Participants (NN = 123) were consecutively recruited from the Brain and Mind Centre in New South Wales, Australia. Internal consistency was determined using Cronbach's alpha. Item-total correlations were evaluated by Pearson's product-moment correlation coefficient. The convergent validity of the DASS-21 was examined by measuring its associations with quality of life and other measures of depression and anxiety. Factorial validity was assessed using confirmatory factor analysis. The DASS-21 demonstrated good internal consistency, adequate convergent validity, and all items exhibited satisfactory item-total correlations. Considering fit indices and factor loadings, the confirmatory factor analysis results provided support for the original 3-factor oblique model consisting of depression, anxiety, and stress factors. The model fit could be further improved with some modifications. Overall, the results indicate that the DASS-21 is a viable self-report screening measure for depression, anxiety, and stress in individuals with ASD and without intellectual disability.

Machine Learning for Differential Diagnosis Between Clinical Conditions With Social Difficulty: Autism Spectrum Disorder, Early Psychosis, and Social Anxiety Disorder.

Differential diagnosis in adult cohorts with social difficulty is confounded by comorbid mental health conditions, common etiologies, and shared phenotypes. Identifying shared and discriminating profiles can facilitate intervention and remediation strategies. The objective of the study was to identify salient features of a composite test battery of cognitive and mood measures using a machine learning paradigm in clinical cohorts with social interaction difficulties. We recruited clinical participants who met standardized diagnostic criteria for autism spectrum disorder (ASD: n = 62), early psychosis (EP: n = 48), or social anxiety disorder (SAD: N = 83) and compared them with a neurotypical comparison group (TYP: N = 43). Using five machine-learning algorithms and repeated cross-validation, we trained and tested classification models using measures of cognitive and executive function, lower- and higher-order social cognition and mood severity. Performance metrics were the area under the curve (AUC) and Brier Scores. Sixteen features successfully differentiated between the groups. The control versus social impairment cohorts (ASD, EP, SAD) were differentiated by social cognition, visuospatial memory and mood measures. Importantly, a distinct profile cluster drawn from social cognition, visual learning, executive function and mood, distinguished the neurodevelopmental cohort (EP and ASD) from the SAD group. The mean AUC range was between 0.891 and 0.916 for social impairment versus control cohorts and, 0.729 to 0.781 for SAD vs neurodevelopmental cohorts. This is the first study that compares an extensive battery of neuropsychological and self-report measures using a machine learning protocol in clinical and neurodevelopmental cohorts characterized by social impairment. Findings are relevant for diagnostic, intervention and remediation strategies for these groups.

Self-reported empathy in adults with autism, early psychosis, and social anxiety disorder.

The Empathy Quotient (EQ) self-report questionnaire is used to measure empathy in individuals with clinical conditions that have been associated with social impairments. In this study, older teens and adults with autism spectrum disorder (ASD; N = 60), early psychosis (EP; N = 51) and social anxiety disorder (SAD; N = 71) and neurotypical controls (NT; N = 26) were compared on the cognitive empathy, emotional reactivity and social skills sub-scales of the Empathy Quotient (EQ) measure. All three clinical groups reported lower cognitive empathy than NT controls, and the ASD group reported lower cognitive empathy than EP and SAD groups. The ASD group reported lower emotional reactivity than the SAD group. All three clinical groups reported lower social skills that NT controls. The poor self-rated empathy for the ASD and EP groups generally reflects previous research that found individuals with these conditions perform relatively poorly on certain objective measures of empathy. However, the poor self-rated cognitive empathy and social skills for the SAD group conflicts with previous research that has found that SAD groups perform well on objective measures of empathy. This suggests that both EQ and objective measures should be used to fully assess empathy in clinical groups.

Validation of the social functioning scale: Comparison and evaluation in early psychosis, autism spectrum disorder and social anxiety disorder.

Social functioning is an important component of mental disorders for assessment and treatment. There is no recognised self-report instrument to measure social functioning across disorders where social impairment is significant. The Social Functioning Scale (SFS) has, however, been used to assess social functioning in psychotic disorders, including Schizophrenia and Early Psychosis. The current study investigated the reliability, validity and sensitivity of the SFS in Early Psychosis, Autism Spectrum Disorder (ASD), Social Anxiety Disorder (SAD) and neurotypical control populations. As expected, all clinical groups showed significant impairment on the total and sub-scale scores of the SFS. The SFS showed good internal consistency and concurrent validity for people diagnosed with SAD and Early Psychosis and a similar factors structure was found for these groups. Participants with ASD reported a relatively low internal consistency and poor concurrent validity, as well as a three-component solution. The SFS has also showed a good sensitivity to separate clinical populations and neurotypical controls. This study supports the use of the SFS for those with SAD and Early Psychosis. Lower internal consistency in ASD populations suggests further research in larger samples is required and that the relationship between its scales are likely different to other populations. Alternative scales or significant other reports may be required for adults with ASD.

Validation of the 36-item and 12-item self-report World Health Organization Disability Assessment Schedule II (WHODAS-II) in individuals with autism spectrum disorder.

The World Health Organization Disability Assessment Schedule II (WHODAS-II) is one of the most widely used generic assessments for measuring disability levels in both clinical and nonclinical populations, with sound psychometrics that is also aligned with the International Classification of Functioning framework. However, its psychometric properties have not been explored extensively in individuals with autism spectrum disorder (ASD). This study examined the psychometric properties of the 36-item and 12-item Self-Report WHODAS-II from 109 individuals diagnosed with ASD and without intellectual disability (IQ ≥ 70). Participants were consecutively recruited from the Brain and Mind Centre in New South Wales, Australia. The WHODAS-II showed adequate internal consistency for all domain scores (α = 0.78-0.97 for 36-item) and for the summary scale (α = 0.95 for 36-item; 0.86 for 12-item). All items also exhibited satisfactory correlations with their respective domain (r = 0.39-0.94 for 36-item) and summary scores (r = 0.42-0.71 for 36-item; 0.42-0.67 for 12-item), except item 4.5 "sexual activity" from the 36-item WHODAS-II (r = 0.19). Concurrent validity was shown by moderate correlations between similar constructs across the WHODAS-II and the World Health Organization Quality of Life BREF (Ps < 0.05). The second-order 7-factor model showed the best fit for the 36-item WHODAS-II, while the second-order 6-factor model demonstrated an acceptable fit for the 12-item WHODAS-II. The model fit could be improved with some modifications. The Schmid-Leiman transformation further confirmed the appropriateness of the second-order factor structure. Overall, the results indicated that the WHODAS-II is a viable generic self-report measure for disability in autistic individuals without ID. Autism Res 2019, 12: 1101-1111. © 2019 International Society for Autism Research, Wiley Periodicals, Inc. LAY SUMMARY: The majority of autistic people have a disability with a profound or severe limitation in their core activities. However, there is currently limited research identifying reliable and valid self-report measures for disability in the autistic population. This study examined the psychometric properties of the World Health Organization Disability Assessment Schedule II (WHODAS-II) from 109 autistic individuals without intellectual disability. Our results suggest that the WHODAS-II is a viable generic self-report measure for disability in autistic individuals.

Disability, functioning, and quality of life among treatment-seeking young autistic adults and its relation to depression, anxiety, and stress.

In this study, we consecutively recruited treatment-seeking young autistic adults without intellectual impairment aged 16-30 years who presented to a mental health service and evaluated general health (distress, quality of life, and disability), functioning (work loss days and social functioning), and mood symptoms (depression, anxiety, and stress) in those diagnosed with autism spectrum disorder (n = 96). This group was compared to young adults presenting to the same service with primary mental health disorders (depression, n = 343; bipolar, n = 132; psychosis, n = 166; and anxiety, n = 303). This study also investigated the influence of mood symptoms on general health and functioning in the autism spectrum disorder group. Young autistic adults reported significant general health and functioning impairments that were of similar degree to those presenting with primary mental health disorders. Interestingly, the autistic group also reported similarly high levels of mood symptoms to those with primary depressive and anxiety disorders. In the autistic group, depressive symptoms were strongly associated with distress, quality of life, and work loss days, while stress symptoms were strongly associated with disability. This study highlights further research, and mental health services are required specifically targeting young autistic adults to address their significant unmet needs.

Mesenchymal stem/stromal cells genetically engineered to produce vascular endothelial growth factor for revascularization in wound healing and ischemic conditions.

Mesenchymal stem/stromal cells (MSCs) may be able to improve ischemic conditions as they can actively seek out areas of low oxygen and secrete proangiogenic factors. In more severe trauma and chronic cases, however, cells alone may not be enough. Therefore, we have combined the stem cell and angiogenic factor approaches to make a more potent therapy. We developed an engineered stem cell therapy product designed to treat critical limb ischemia that could also be used in trauma-induced scarring and fibrosis where additional collateral blood flow is needed following damage to and blockage of the primary vessels. We used MSCs from normal human donor marrow and engineered them to produce high levels of the angiogenic factor vascular endothelial growth factor (VEGF). The MSC/VEGF product has been successfully developed and characterized using good manufacturing practice (GMP)-compliant methods, and we have completed experiments showing that MSC/VEGF significantly increased blood flow in the ischemic limb of immune deficient mice, compared to the saline controls in each study. We also performed safety studies demonstrating that the injected product does not cause harm and that the cells remain around the injection site for more than 1 month after hypoxic preconditioning. An on-demand formulation system for delivery of the product to clinical sites that lack cell processing facilities is in development.

Autism, Early Psychosis, and Social Anxiety Disorder: a transdiagnostic examination of executive function cognitive circuitry and contribution to disability.

The disability burden in clinical cohorts with social impairment is significant, leading to poor functional outcomes. Some of this impairment has been linked to executive dysfunction. In this study, a transdiagnostic approach was taken to identify executive function (EF) processes in young adults that may underpin social impairment and to evaluate their contribution to disability. Comparisons were made between three prominent disorders that are characterized by social impairments, Autism Spectrum Disorder (ASD), Early Psychosis (EP) and Social Anxiety Disorder (SAD), as well as a neurotypically developing group (TYP). We examined whether overall disability could be predicted by neuropsychological and self-report assessments of EF. Our study showed that ASD participants demonstrated impaired performance on most domains of EF compared to the TYP group (mental flexibility, sustained attention and fluency) while the EP group showed impairment on sustained attention and attentional shifting. The SAD participants showed EF impairment on self-report ratings, even though their objective performance was intact. Self-reports of EF explained a significant percentage (17%) of disability in addition to the variance explained by other predictors, and this was particularly important for ASD. This is the first study to compare EF measures across clinical groups of social impairment and suggests unique cognitive-circuitry that underpins disability within groups. Impairments in EF were broad in ASD and predicted disability, EP impairments were specific to attentional processes and SAD impairments likely relate to negative self-monitoring. Self-report, as opposed to performance-based EF, provided best capacity to predict disability. These findings contribute to transdiagnostic circuitry models and intervention strategies.

Preclinical evaluation of mesenchymal stem cells overexpressing VEGF to treat critical limb ischemia.

Numerous clinical trials are utilizing mesenchymal stem cells (MSC) to treat critical limb ischemia, primarily for their ability to secrete signals that promote revascularization. These cells have demonstrated clinical safety, but their efficacy has been limited, possibly because these paracrine signals are secreted at subtherapeutic levels. In these studies the combination of cell and gene therapy was evaluated by engineering MSC with a lentivirus to overexpress vascular endothelial growth factor (VEGF). To achieve clinical compliance, the number of viral insertions was limited to 1-2 copies/cell and a constitutive promoter with demonstrated clinical safety was used. MSC/VEGF showed statistically significant increases in blood flow restoration as compared with sham controls, and more consistent improvements as compared with nontransduced MSC. Safety of MSC/VEGF was assessed in terms of genomic stability, rule-out tumorigenicity, and absence of edema or hemangiomas in vivo. In terms of retention, injected MSC/VEGF showed a steady decline over time, with a very small fraction of MSC/VEGF remaining for up to 4.5 months. Additional safety studies completed include absence of replication competent lentivirus, sterility tests, and absence of VSV-G viral envelope coding plasmid. These preclinical studies are directed toward a planned phase 1 clinical trial to treat critical limb ischemia.

Human Mesenchymal Stem Cells Genetically Engineered to Overexpress Brain-derived Neurotrophic Factor Improve Outcomes in Huntington's Disease Mouse Models.

Huntington's disease (HD) is a fatal degenerative autosomal dominant neuropsychiatric disease that causes neuronal death and is characterized by progressive striatal and then widespread brain atrophy. Brain-derived neurotrophic factor (BDNF) is a lead candidate for the treatment of HD, as it has been shown to prevent cell death and to stimulate the growth and migration of new neurons in the brain in transgenic mouse models. BDNF levels are reduced in HD postmortem human brain. Previous studies have shown efficacy of mesenchymal stem/stromal cells (MSC)/BDNF using murine MSCs, and the present study used human MSCs to advance the therapeutic potential of the MSC/BDNF platform for clinical application. Double-blinded studies were performed to examine the effects of intrastriatally transplanted human MSC/BDNF on disease progression in two strains of immune-suppressed HD transgenic mice: YAC128 and R6/2. MSC/BDNF treatment decreased striatal atrophy in YAC128 mice. MSC/BDNF treatment also significantly reduced anxiety as measured in the open-field assay. Both MSC and MSC/BDNF treatments induced a significant increase in neurogenesis-like activity in R6/2 mice. MSC/BDNF treatment also increased the mean lifespan of the R6/2 mice. Our genetically modified MSC/BDNF cells set a precedent for stem cell-based neurotherapeutics and could potentially be modified for other neurodegenerative disorders such as amyotrophic lateral sclerosis, Alzheimer's disease, and some forms of Parkinson's disease. These cells provide a platform delivery system for future studies involving corrective gene-editing strategies.

Human Myoblast and Mesenchymal Stem Cell Interactions Visualized by Videomicroscopy.

Muscle-derived progenitor cell (myoblast) therapy has promise for the treatment of denervated, weakened, and fibrotic muscle. The best methods for injecting myoblasts to promote fusion and retention have yet to be determined, however. Mesenchymal stem/stromal cells have also been reported to have beneficial effects in restoring damaged tissue, through increasing vascularization and reducing inflammation. The interactions between human primary skeletal myoblasts and bone marrow-derived mesenchymal stem/stromal cells were examined using time-lapse images put into video format. Of interest, there is a high degree of cell-to-cell interaction with microparticles transferring between both cell types, and formation of nanotubules to bridge cytoplasmic contents between the two types of cell. This model provides an in vitro platform for examining mechanisms for cell-to-cell interaction preceding myoblast fusion.

Sexual health of Indian immigrant men in Australia: an exploratory research on help-seeking attitudes.

The help-seeking attitudes for sexual health of Indian men living in Australia was explored. Of all survey respondents (n=225), many preferred to seek help from medical doctors. Young (18-25 years) Indian men were three times more likely to prefer a specialist medical doctor than older men. Ethnicity and gender of the medical doctor was 'not important' for the majority of men. Most men preferred to seek help from their regular general practitioner.

Prevalence and risk factors for musculoskeletal symptoms with computer based work across occupations.

OBJECTIVES: The purpose of this study was to compare the 12-month prevalence of musculoskeletal symptoms and risk factors associated with computer based work between occupations in a sample of Australian public sector employees. METHOD: A cross-sectional study was completed with employees of 6 government departments. An online survey was electronically distributed to over 8,000 employees characterised by a range of occupational groups and levels of employment. Data collected included individual and employment characteristics, estimation of hours worked with a computer per day and self-reported musculoskeletal symptoms in the upper extremity and spinal areas using the Nordic Musculoskeletal Questionnaire. RESULTS: Responses from 934 completed surveys could be used. There was no significant difference in the prevalence of reported musculoskeletal symptoms between occupational groups except for the wrist/hand and elbow areas. Estimated duration of computer work per day was significantly associated with increased musculoskeletal symptoms in the neck (OR 1.41, 95%CI: 1.09 to 1.83), wrist/hand/s (OR 1.46, 95%CI: 1.17 to 1.83) and elbow/s (OR 1.41, 95%CI: 1.07 to 1.85) areas, with the finding of a linear relationship between hours worked and prevalence of symptoms. A greater proportion of employees in higher level management and professional occupational groups were found to be working with a computer in excess of 6 to 8 hours per day compared with those in non-professional (administration and secretarial) groups. CONCLUSION: Hours worked with a computer per day was a significant risk factor for reported musculoskeletal symptoms amongst all occupational groups working in Australian public sector offices. No significant difference in the level of risk was found between occupations.

Examination of mesenchymal stem cell-mediated RNAi transfer to Huntington's disease affected neuronal cells for reduction of huntingtin.

Huntington's disease (HD) is a fatal, autosomal dominant neurodegenerative disorder caused by an expanded trinucleotide (CAG) repeat in exon 1 of the huntingtin gene (Htt). This expansion creates a toxic polyglutamine tract in the huntingtin protein (HTT). Currently, there is no treatment for either the progression or prevention of the disease. RNA interference (RNAi) technology has shown promise in transgenic mouse models of HD by reducing expression of mutant HTT and slowing disease progression. The advancement of RNAi therapies to human clinical trials is hampered by problems delivering RNAi to affected neurons in a robust and sustainable manner. Mesenchymal stem cells (MSC) have demonstrated a strong safety profile in both completed and numerous ongoing clinical trials. MSC exhibit a number of innate therapeutic effects, such as immune system modulation, homing to injury, and cytokine release into damaged microenvironments. The ability of MSC to transfer larger molecules and even organelles suggested their potential usefulness as delivery vehicles for therapeutic RNA inhibition. In a series of model systems we have found evidence that MSC can transfer RNAi targeting both reporter genes and mutant huntingtin in neural cell lines. MSC expressing shRNA antisense to GFP were found to decrease expression of GFP in SH-SY5Y cells after co-culture when assayed by flow cytometry. Additionally MSC expressing shRNA antisense to HTT were able to decrease levels of mutant HTT expressed in both U87 and SH-SY5Y target cells when assayed by Western blot and densitometry. These results are encouraging for expanding the therapeutic abilities of both RNAi and MSC for future treatments of Huntington's disease.

Characterization and in vivo testing of mesenchymal stem cells derived from human embryonic stem cells.

Mesenchymal stem cells (MSCs) have been shown to contribute to the recovery of tissues through homing to injured areas, especially to hypoxic, apoptotic, or inflamed areas and releasing factors that hasten endogenous repair. In some cases genetic engineering of the MSC is desired, since they are excellent delivery vehicles. We have derived MSCs from the human embryonic stem cell (hESC) line H9 (H9-MSCs). They expressed CD105, CD90, CD73, and CD146, and lacked expression of CD45, CD34, CD14, CD31, and HLA-DR, the hESC pluripotency markers SSEA-4 and Tra-1-81, and the hESC early differentiation marker SSEA-1. Marrow-derived MSCs showed a similar phenotype. H9-MSCs did not form teratoma in our initial studies, whereas the parent H9 line did so robustly. H9-MSCs differentiated into bone, cartilage, and adipocytes in vitro, and displayed increased migration under hypoxic conditions. Finally, using a hindlimb ischemia model, H9-MSCs were shown to home to the hypoxic muscle, but not the contralateral limb, by 48 h after IV injection. In summary, we have defined methods for differentiation of hESCs into MSCs and have defined their characteristics and in vivo migratory properties.