Family perspectives of how their relatives with mental illness benefit from Clubhouse participation: a qualitative inquiry.

BACKGROUND: Although researchers have demonstrated the benefits of psychosocial Clubhouse participation on a number of clinical and psychosocial outcomes, few studies have investigated the consumer's participation from the perspectives of others. AIM: This study aimed to investigate family members' perspectives of how Clubhouse programming has affected consumers' recovery. METHOD: Twenty-four relatives of Clubhouse members were interviewed using a semi-structured protocol. Relatives were nominated by their Clubhouse members as their family member who provided them with the most social support. All interviews were transcribed and underwent content analysis yielding multilevel coding. RESULTS: Four main dimensions emerged from family interviews about how Clubhouses affected their relatives. These dimensions aligned with areas of clinical recovery and personal growth. Family members observed and noted changes in: (1) positive affective changes, (2) improved goal directed and challenging behaviors, (3) positive attitude changes and (4) greater social interactions. CONCLUSION: As one of the first studies to document the perspectives of the relatives of Clubhouse members, this exploratory study indicates that family members recognize positive changes in their Clubhouse family members and these changes align with areas of functional recovery. Implications for practice and future studies are discussed.

Daisy Chain Rotaxanes Made from Interlocked DNA Nanostructures.

We report the stepwise assembly of supramolecular daisy chain rotaxanes (DCR) made of double-stranded DNA: Small dsDNA macrocycles bearing an axle assemble into a pseudo-DCR precursor that was connected to rigid DNA stoppers to form DCR with the macrocycles hybridized to the axles. In presence of release oligodeoxynucleotides (rODNs), the macrocycles are released from their respective hybridization sites on the axles, leading to stable mechanically interlocked DCRs. Besides the expected threaded DCRs, certain amounts of externally hybridized structures were observed, which dissociate into dumbbell structures in presence of rODNs. We show that the genuine DCRs have significantly higher degrees of freedom in their movement along the thread axle than the hybridized DCR precursors. Interlocking of DNA in DCRs might serve as a versatile principle for constructing functional DNA nanostructures where the movement of the subunits is restricted within precisely confined tolerance ranges.

Comparison of the responses of different recombinant fish type I interferons against betanodavirus infection in grouper.

The nervous necrosis virus (NNV) is an aquatic virus that can infect more than 30 species including the grouper, which is a valuable fish species in Taiwan. NNV causes up to 90-100% mortality in the aquaculture industry. Interferons (IFNs) are a family of cytokines that stimulate the expression of numerous proteins to protect the host against viruses and possess very unique specific characteristics in fish. The cross-reactivity of heterologous IFNs on grouper cells and larvae has not been well-studied to date. To evaluate and compare the anti-NNV effect of different fish IFNs in grouper, we successfully synthesized, subcloned, expressed and purified several fish type I IFNs in the present study: grouper (gIFN), salmon (sIFN), seabass (sbIFN) and tilapia (tpIFN). The gIFN and sIFN proteins up-regulated myxovirus resistance protein (Mx) gene expression in grouper kidney (GK) cells, but similar effects were not observed for sbIFN and tpIFN. Following co- and pre-treatment with the 4 types of IFNs with NNV infection in GK cells, sIFN exhibited the strongest antiviral ability to suppress NNV gene replication (especially at 24 h) and significantly reduced the cytopathic effect (CPE) at 72 h, followed by gIFN. Unsurprisingly, sbIFN and tpIFN had no significant effect on CPE but slightly suppressed NNV gene replication. The cytotoxicity of these four fish IFNs on GK cells was also examined for the first time. In the in vivo test, we confirmed that gIFN and sIFN had a significant protective effect against NNV when administered by intraperitoneal (IP) injection and the oral route in Malabar grouper (Epinephelus malabaricus) larvae. This study compared the protective effects of IFNs from various fish species against NNV and demonstrated crosstalk between sIFN and grouper cells for the first time. These results provide information concerning the efficacy of fish IFNs for possible therapeutic applications.

Mechanisms relevant to the enhanced virulence of a dihydroxynaphthalene-melanin metabolically engineered entomopathogen.

The entomopathogenic fungus Metarhizium anisopliae MA05-169 is a transformant strain that has been metabolically engineered to express dihydroxynaphthalene-melanin biosynthesis genes. In contrast to the wild type strain, the transformant displays a greater resistance to environmental stress and a higher virulence toward target insect host. However, the underlying mechanisms for these characteristics remain unclear; hence experiments were initiated to explore the possible mechanism(s) through physiological and molecular approaches. Although both transformant and wild type strains could infect and share the same insect host range, the former germinated faster and produced more appressoria than the latter, both in vivo and in vitro. The transformant showed a significantly shorter median lethal time (LT50) when infecting the diamondback moth (Plutella xylostella) and the striped flea beetle (Phyllotreta striolata), than the wild type. Additionally, the transformant was more tolerant to reactive oxygen species (ROS), produced 40-fold more orthosporin and notably overexpressed the transcripts of the pathogenicity-relevant hydrolytic enzymes (chitinase, protease, and phospholipase) genes in vivo. In contrast, appressorium turgor pressure and destruxin A content were slightly decreased compared to the wild type. The transformant's high anti-stress tolerance, its high virulence against five important insect pests (cowpea aphid Aphis craccivora, diamondback moth Pl. xylostella, striped flea beetle Ph. striolata, and silverleaf whitefly Bemisia argentifolii) and its capacity to colonize the root system are key properties for its potential bio-control field application.

Predictors of loneliness of clubhouse members.

OBJECTIVE: Loneliness can impede subjective experiences of recovery. This study examines the relationship between clubhouse participation and loneliness using standardized instruments while controlling for age, gender, living status, and social network characteristics. METHOD: A random sample of 126 members from one clubhouse was recruited for this cross-sectional investigation. A hierarchical multiple regression analysis was performed to examine the association between participation and loneliness. RESULTS: A greater number of clubhouse visits, greater perceived availability of social support and higher levels of overall satisfaction with social network relationships were significantly related to a lower level of loneliness after controlling for covariates. CONCLUSIONS AND IMPLICATIONS FOR PRACTICE: Service providers may want to pay more attention to those members experiencing loneliness and help them connect with others. Future studies using longitudinal analyses are needed to further assess the causal relationship between clubhouse participation and loneliness. Multiple aspects of participation should be examined.

Association of aberrant expression of sex-determining gene fibroblast growth factor 9 with Sertoli cell-only syndrome.

OBJECTIVE: To investigate the expressions of fibroblast growth factor 9 (FGF9) in normal testes and in testes with Sertoli cell-only syndrome (SCOS), explore the biological function of testicular FGF9, and identify the sequence variants of FGF9 gene in patients with SCOS. DESIGN: Retrospective case study. SETTING: University reproductive clinic. PATIENT(S): Forty-one patients with SCOS, seven with normal spermatogenesis, and 100 controls. INTERVENTION(S): Protein expressions of testicular FGF9 and sequence variants of FGF9 gene in normal controls and patients with SCOS were studied. The biological function and regulation of testicular FGF9 were assessed in vitro. MAIN OUTCOME MEASURE(S): Expression profiles of testicular FGF9, effects of FGF9 on germ cell proliferation, and sequence variants of the FGF9 gene. RESULT(S): FGF9 was predominately expressed in the cytoplasm of Leydig cells of normal testis; its expression was significantly decreased in patients with SCOS. Conditioned medium of FGF9-treated Leydig cells stimulated germ cell proliferation. A promoter polymorphism (c.-712C→T) of the FGF9 gene attenuated the promoter activity, which contributes to one of the causes of its low expression. CONCLUSION(S): In addition to the role of sex determination, FGF9 is expressed in postnatal Leydig cells and is involved in cell-to-cell interaction of testicular function. Aberrant expression of testicular FGF9 is associated with SCOS.

Characterization of 3-hydroxyisobutyrate dehydrogenase, HIBADH, as a sperm-motility marker.

PURPOSE: Asthenozoospermia is a major cause of male infertility. However, the molecular mechanisms underlying sperm-motility defects remain largely unknown in the majority of cases. In our previous study, we applied a proteomic approach to identify unknown proteins that were downregulated in spermatozoa with low motility compared to spermatozoa with good motility. Several sperm motility- related proteins have been identified. In this study, 3-hydroxyisobutyrate dehydrogenase (HIBADH), one of the proteins identified using the proteomic tools, is further characterized. METHODS: Reverse-transcription polymerase chain reactions (RT-PCR), western blotting, and immunofluorescence assays (IFA) were preformed to investigate the expression pattern. The enzymatic activity of HIBADH was evaluated in sperm with good (>50 %), moderate (< 50 %) and lower motility (< 20 %). RESULTS: Using RT-PCR, we found that transcripts of HIBADH are enriched in the cerebellum, heart, skeletal muscle, uterus, placenta, and testes of male humans. In western blotting, it is expressed in the placenta, testes, and spermatozoa. During spermiogenesis, HIBADH is located at the mid-piece (a specialized development from the mitochondria) of elongating, elongated, and mature sperm. The enzymatic activity of HIBADH in sperm with moderate and lower motility were significantly reduced compared with good motility (P<0.0001 and P<0.05, respectively). CONCLUSIONS: Our study indicated that HIBADH is involved in the mitochondrial function of spermatozoa, and maintains sperm motility. It may serve as a sperm-motility marker.

Protein tyrosine phosphatase non-receptor type 14 is a novel sperm-motility biomarker.

PURPOSE: To understand the molecular basis of sperm-motility and to identify related novel motility biomarkers. METHODS: Two-dimensional electrophoresis (2DE) followed by Reverse-phase-nano-high-performance liquid chromatography-electrospray ionization tandem mass spectrometry (RP-nano-HPLC-ESI-MS/MS) were applied to establish the human sperm proteome. Then the sperm proteome of moderate-motile human sperm fraction and that of good-motile human sperm fraction from pooled spermatozoa of forty normozoospermic donors (Group 1 subjects) were compared to identify the dysregulated proteins. Among these down-regulated proteins, Protein tyrosine phosphatase non-receptor type 14 (PTPN14) was chosen to reconfirm by Western blotting and semi-quantitative reverse transcription polymerase chain reaction. For clinical application, Western blotting and real-time reverse transcription polymerase chain reaction was performed to compare the expression level of PTPN14 in (Group 2 subjects) nine normozoospermic controls and thirty-three asthenozoospermic patients (including 21 mild asthenozoospermic cases and 12 severe cases). Finally, bioinformatic tools prediction and immunofluorescence assay were performed to elucidate the potential localization of PTPN14. RESULTS: The expression levels of three proteins were observed to be lower in the moderate-motile sperm fraction than in good-motile sperm of group 1 subjects. Among three proteins with persistent down-regulation in the moderate-motile sperm, we reconfirmed that the expression level of PTPN14 was significantly lower in both mRNA and protein levels from the moderate-motile sperm fraction. Further, down-regulation of PTPN14 was found at the translational and transcriptional level in the asthenozoospermic men. Finally, Bioinformatic tools prediction and immunofluorescence assay showed that PTPN14 maybe predominantly localized at the mitochondria in the midpiece of human ejaculated sperm. CONCLUSIONS: Proteomics tools were applied to identify three possible sperm motility-related proteins. Among these proteins, PTPN14 was highly likely a novel sperm-motility biomarker and a potential mitochondrial protein.

Hierarchical functionalisation of single-wall carbon nanotubes with DNA through positively charged pyrene.

A simple and efficient method to link reversibly DNA to SWNTs via electrostatic interaction is reported. The DNA/nanotube hybrids are characterised by a combination of gel electrophoresis and AFM.

Fibroblast growth factor 9 stimulates steroidogenesis in postnatal Leydig cells.

Fibroblast growth factor 9 (FGF9) is a potent mitogen and survival factor required for morphogenesis during embryonic development and numerous biological functions at adulthood. The reproductive phenotype of mice lacking Fgf9 gene exhibits male to female sexual reversal, suggesting a crucial role of Fgf9 in male sex determination. Our previous study showed that polymorphic microsatellite of FGF9 genes is associated with 46XY female with ambiguous genitalia, implying that the aberrant expression of FGF9 might affect androgen secretion. In this study, we aimed to investigate the effect of FGF9 on testosterone production in mouse Leydig cell and to study the signalling pathways by which FGF9 modulate steroidogenesis. Our results show that mRNAs of Fgf9 and Fgfr isoforms (Fgfr2IIIc, Fgfr3 and Fgfr4) were all expressed in mouse Leydig cells. FGF9 significantly stimulates mouse Leydig cell testosterone production in a dose- and time-dependent manner. Ras-MAPK, PI3K and PKA signalling pathways are involved in the FGF9-induced steroidogenesis. These results provide supportive evidence linking the aberrant expression of FGF9 to human gonadal dysgenesis and suggest a role of FGF9 in postnatal testicular development.

Direct conductance measurements of short single DNA molecules in dry conditions.

We present a study of electronic transport in short (12-base-pair) DNA duplexes covalently bonded (via thiol groups) to two gold electrodes obtained by a mechanically controllable break junction (MCJB) technique in dry conditions. A large number of DNA junctions have been repeatedly formed in order to obtain a conductance histogram that reveals a peak which corresponds to the conductance of a single DNA molecule. We observed that the conductivity of a DNA increases upon increasing the content of G:C base pairs in the duplex. With our method we are able to obtain a reliable value of a single DNA conductance and subsequently measure its current-voltage (I-V) characteristics. In contrast to the electronic transport measurements performed with long DNA sequences (hundreds of base pairs) where the obtained conductance values vary a lot with environmental conditions, our values obtained for the short DNA sequences are consistent with the values reported for comparable sequences in aqueous solution.

Posttranscriptional regulation of CDC25A by BOLL is a conserved fertility mechanism essential for human spermatogenesis.

CONTEXT: Human BOLL is known as a meiotic regulator, and CDC25A is considered as a potential RNA target of BOLL. OBJECTIVE: The aim of the study was to clarify the relationship between BOLL and CDC25A expressions in human testes and to explore the mechanism by which BOLL regulates CDC25A. DESIGN AND SETTING: A prospective experimental study was conducted at a university-based medical center. PARTICIPANTS AND INTERVENTIONS: BOLL protein, CDC25A mRNA, and CDC25A protein expressions, as well as spermatocyte numbers in the testes of 32 infertile men were measured. The interaction between BOLL protein and CDC25A mRNA was assessed in in vitro studies. MAIN OUTCOME MEASURES: Protein and RNA expressions, relationships between expression profiles, CDC25A mRNA binding site for BOLL, and the effects of BOLL on CDC25A mRNA stability and translatability were measured. RESULTS: The protein expressions of BOLL and CDC25A are significantly decreased in patients with spermatogenic failure, with the lowest levels detected in patients with meiotic arrest. Both protein expressions are significantly correlated with spermatocyte numbers. Expressional profiling analysis among BOLL protein, CDC25A mRNA, and CDC25A protein suggests a causal relationship between BOLL and CDC25A. BOLL specifically binds to a 21-nucleotide region of the CDC25A 3'UTR, and this region is evolutionarily conserved. A U-rich region within this 21-nucleotide sequence is crucial for binding. BOLL stimulates CDC25A translation, and this effect does not involve alteration of mRNA stability. CONCLUSION: CDC25A is subject to translational control by BOLL, which is an evolutionarily conserved mechanism. A decreased CDC25A expression caused by lack or decrease of BOLL may be associated with male infertility.

A three-branched DNA template for carbon nanotube self-assembly into nanodevice configuration.

We report here the first realization of an artificial branched DNA template where a single wall carbon nanotube is positioned with the necessary geometry of an individually gated field effect transistor.

DAZL protein expression in mouse preimplantation embryo.

OBJECTIVE: To evaluate the expression pattern of Dazl (deleted in azoospermia-like) protein in the mouse preimplantation embryo. DESIGN: Experimental study. SETTING: Medical research laboratory in a university hospital. ANIMAL(S): Twenty female 28- to 35-day-old FVB mice. INTERVENTION(S): Embryo collection at 1.5, 2.5, and 3.5 days postcoitus (plug date, 0.5 d postcoitus) to examine the Dazl protein expression from the two-cell embryo to the blastocyst. MAIN OUTCOME MEASURE(S): Dazl protein expression was analyzed by immunofluorescent staining. RESULT(S): There is abundant expression of Dazl protein in the cytoplasm of the blastomere. Strong fluorescent signals of Dazl protein expression were found in preimplantation embryo cytoplasm, including two-cell, eight-cell, morula, and blastocyst. CONCLUSION(S): By using an antibody raised against mouse Daz-like protein (Dazl), we showed that Dazl protein is present in all cleaving stages of the preimplantation embryo. This is the first report on the protein expression of a Dazl gene during embryogenesis in mice. However, further study is needed to evaluate the molecular functional role of Dazl.

Expression of various CDC25B isoforms in human spermatozoa.

OBJECTIVE: To explore the role of CDC25B protein in postmeiotic germ cells. DESIGN: In vitro experiment. SETTING: University-based reproductive genetics laboratory. PATIENT(S): Fertile and infertile volunteers. INTERVENTION(S): Reverse transcription-polymerase chain reaction (RT-PCR), real-time RT-PCR, and immunostaining for CDC25B. MAIN OUTCOME MEASURE: Expression profiling of CDC25B in human spermatozoa. RESULT(S): Four splicing variants (CDC25B1, B2, B3, and B4) are expressed in human spermatozoa. Immunofluorescence staining showed strong homogeneous staining in the midpiece of spermatozoa, and weak staining in the principal piece and cytosol of the head. The messenger RNA (mRNA) transcript level of CDC25B was increased in sperm samples of men with asthenospermia. CONCLUSION(S): The expression of CDC25B in different cellular compartments of human spermatozoa suggests that there are diverse noncell-cycle-related functions of CDC25B in terminally differentiated human germ cells.

Association of DAZL haplotypes with spermatogenic failure in infertile men.

OBJECTIVE: To identify novel DAZL single nucleotide polymorphisms (SNPs) and to compare allele/genotype frequencies, linkage disequilibrium (LD) characteristics, and DAZL haplotypes between fertile and infertile men. DESIGN: Prospective case study. SETTING: University genetics laboratory and reproductive clinics. PATIENT(S): Two hundred thirty-one infertile men and 191 men with proven fertility. INTERVENTION(S): Single strand conformation polymorphism and sequence analysis for DAZL gene polymorphism screening were done for all subjects enrolled. MAIN OUTCOME MEASURE(S): Novel SNPs, allele/genotype frequencies, LD characteristics, and DAZL haplotypes between fertile and infertile men. RESULT(S): Five SNPs were identified: 260A>G, 386A>G, 520+34c>a, 584+28c>t and 796+36g>a. SNP 386A>G was significantly associated with spermatogenic failure and was mainly heterozygous in infertile patients. The major haplotypes in infertile men were AACTA (45.8%), followed by AACCG, AAATA, AAACG, and GGACG for 260A>G/386A>G/520+34c>a/584+28c>t/796+36g>a. The major haplotypes for the control subjects were AACCG (41.7 %), AAATA, and AACTA. Of all haplotypes, five showed significant differences in frequency between infertile men and control subjects. Haplotypes AACTA, AAACG, and GGACG were overtransmitted in patients with spermatogenic failure, whereas haplotypes AACCG and AAATA were undertransmitted in these patients. CONCLUSION(S): Our study suggests the association of autosomal DAZL haplotypes with human spermatogenic failure.

Association of spermatogenic failure with decreased CDC25A expression in infertile men.

BACKGROUND: DAZ gene family is crucial for human spermatogenesis that requires the precise co-ordination of cell cycle events. CDC25A is recognized as the downstream substrate of DAZ gene family and is thought to function on the M-phase regulation of cell cycles. We investigated the expression profiles of CDC25A in the testes of infertile men and evaluated the relationship between CDC25A levels and testicular phenotype, clinical hormonal parameters and sperm retrieval results. METHODS: The protein and mRNA transcript levels of CDC25A in the testes of 40 azoospermic men were determined by immunohistochemistry and quantitative real-time-PCR. CDC25A in human spermatozoa was investigated by western blotting and immunofluorescence staining. RESULTS: The CDC25A protein was expressed mainly in spermatocyte, spermatid and spermatozoa. CDC25A transcript levels were significantly decreased (P = 0.0009) in patients with spermatogenic failure, especially in men with meiotic arrest and Sertoli cell-only syndrome. Significantly higher CDC25A transcript levels were detected in patients with successful sperm retrieval than in patients with failed sperm retrieval (P = 0.005). CONCLUSIONS: Decreased CDC25A is associated with spermatogenic failure and failed sperm retrieval in infertile men. Further studies are necessary to explore the functional roles of CDC25A in human spermatozoa.

Decreased mRNA transcripts of M-phase promoting factor and its regulators in the testes of infertile men.

BACKGROUND: M-phase promoting factor (MPF), which is comprised of Cyclin B and a catalytic subunit, Cdc2, is a key enzyme required for cells to enter M phase in both mitosis and meiosis. MPF activity is controlled by the stimulatory dephosphorylation of the Cdc25 family and the inhibitory phosphorylation of Wee1. We determined the levels of mRNA transcripts of MPF and its regulators in the testes of infertile men, and evaluated the relationship between the transcript levels and patients' testicular phenotypes and sperm retrieval results. METHODS AND RESULTS: The mRNA transcript levels of CDC2, CCNB1, CCNB2, CDC25A, CDC25B, CDC25C and WEE1 in the testes of 37 azoospermic patients were examined by quantitative real-time polymerase chain reaction. Significant decreases in CDC2, CCNB1, CCNB2, CDC25A, CDC25C and WEE1 mRNA transcript levels were detected in patients with spermatogenic failure. CDC2 mRNA transcript levels correlated significantly with those of CCNB1 and CCNB2 mRNA. Significantly higher CDC2, CCNB1, CCNB2, CDC25C and WEE1 mRNA transcript levels were detected in 18 patients with successful sperm retrieval than in 11 patients with failed sperm retrieval. CONCLUSIONS: We suggest that the decreased mRNA transcripts of MPF and its regulators play important roles in human spermatogenesis.