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The Y-chromosomal gene SRY acts as the primary trigger for male sex determination in mammalian embryos. Correct regulation of SRY is critical: aberrant timing or level of Sry expression is known to disrupt testis development in mice and we hypothesize that mutations that affect regulation of human SRY may account for some of the many cases of XY gonadal dysgenesis that currently remain unexplained. However, the cis-sequences involved in regulation of Sry have not been identified, precluding a test of this hypothesis. Here, we used a transgenic mouse approach aimed at identifying mouse Sry 5′ flanking regulatory sequences within 8 kb of the Sry transcription start site (TSS). To avoid problems associated with conventional pronuclear injection of transgenes, we used a published strategy designed to yield single-copy transgene integration at a defined, transcriptionally open, autosomal locus, Col1a1. None of the Sry transgenes tested was expressed at levels compatible with activation of Sox9 or XX sex reversal. Our findings indicate either that the Col1a1 locus does not provide an appropriate context for the correct expression of Sry transgenes, or that the cis-sequences required for Sry expression in the developing gonads lie beyond 8 kb 5′ of the TSS. [ABSTRACT FROM AUTHOR]

Background: During the COVID-19 pandemic, educational interventions have become necessary to prevent the spread of health-related misinformation among Hong Kong older adults. The primary objective of this study was to assess the feasibility of a student-led, telephone-delivered intervention to improve COVID-19-related health knowledge among Hong Kong older adults. The secondary objective was to evaluate the impact of the intervention on the student volunteers., Methods: Twenty-five participants aged 65 or above who were able to communicate in Cantonese and had no hearing or cognitive impairments were recruited for this longitudinal pre-post-study from a community center in Hong Kong. The pilot telephone-delivered intervention consisted of five telephone call sessions conducted by 25 student volunteers. Each participant was paired with the same volunteer throughout the intervention. The first four sessions included pre-tests that assessed the participants' understanding of three COVID-19-related themes: medication safety, healthcare voucher scheme, and COVID-19 myth-busting. Standardized explanations of the pre-test questions were offered to participants during the phone calls. In the last session, a post-test on all the themes was conducted. The intervention's feasibility was assessed based on (a) percentage changes in the participants' test scores, (b) attrition rate, and (c) the acceptability of the intervention by the participants. The impact of the intervention on the student volunteers was evaluated based on a student feedback survey. There was no control group., Results: Significant improvements in the participants' test scores (out of 100%) for all themes were observed after the intervention: from 76 to 95.2% for medication safety, from 64.0 to 88.8% for the healthcare voucher scheme, and from 78.0 to 93.2% for COVID-19 myth-busting. The average improvement in test scores of the three themes was 18.4% (95% CI 12.2 to 24.6%). Most participants were satisfied with the program. The student feedback survey suggested that the intervention enhanced students' communication skills and understanding of Hong Kong older adults., Conclusion: This pilot study offers initial evidence of the potential and feasibility of student-led, telephone-delivered educational interventions for the transfer of COVID-19-related knowledge to older adults and their benefits for the student volunteers. Future studies should include larger samples and a control group., (© 2022. The Author(s).)

Given that social withdrawal has been consistently linked to intimacy, an examination of the mediating effect of interpersonal distrust on the relationship between social withdrawal and intimacy could offer potential insights into a possible explanatory mechanism. Therefore, the purpose of this study was to investigate interpersonal distrust as a mediator of the relationship between social withdrawal and intimacy, and whether retrospective bullying victimization moderates this mediation. Participants included 160 university students who completed measures of social withdrawal, interpersonal distrust, intimacy, and retrospective bullying victimization. Results indicated that interpersonal distrust mediated the relationship between social withdrawal and intimacy. Retrospective bullying victimization in primary school, however, did not moderate the mediation model. In light of these findings, appropriate implications and suggestions for interventions are discussed.

Background: Over the last decade, neoadjuvant systemic therapy (NAST) has gained considerable popularity and its use has been extended to include breast cancer patients with operable node-positive disease. It may no longer be necessary to commit patients who are node-positive at presentation to axillary dissection if they become clinically node-negative after completing NAST. Targeted axillary dissection (TAD) is a technique where the marked pre-NAST positive node is excised along with the sentinel nodes and its response to chemotherapy is assessed and thus helps guide further treatment to the axilla., Methods: The aim of this study was to determine the feasibility of marking positive axillary nodes with a clip and removing the clipped node after neoadjuvant treatment. We also assessed the concordance of the sentinel node with the clipped node., Results: We prospectively evaluated 37 clinically and/or radiologically node-positive patients who underwent NAST. The overall identification rate of the clipped node was 78%. The identification rate was 100% if the clipped node was localized preoperatively and was much lower at 68% in patients who did not have the clipped node localized. The clipped node was not retrieved as the sentinel node in 14% of patients., Conclusion: We present the first Australian series on the feasibility of TAD. TAD is a feasible option in patients having NAST and with every new technique there is a learning curve. With the increasing experience globally and the refinement in marking and localization techniques, the accuracy of performing TAD will likely continue to improve., (© 2019 Royal Australasian College of Surgeons.)

Introduction: The aim of the present study was to evaluate the understanding of generic substitution among health care professionals and members of the general public ("general public") in Hong Kong., Methods: This cross-sectional descriptive study was performed by using a self-completed anonymous questionnaire from March 2015 to May 2017. The questionnaire included demographic data, knowledge of generic drugs, experiences of generic substitution, and views on policy., Results: A total of 2106 general public, 73 doctors, 22 nurses, and 50 pharmacists responded the questionnaire. In all, 41.2% of the general public was aware that generic drugs have the same active ingredients. Although a majority of the health care professionals knew that generic drugs have the same active ingredients (doctors: 79.5%; nurses: 86.4%; pharmacists: 98.0%), many were unaware of bioequivalence (doctors: 37.0%; nurses: 18.2%; pharmacists: 50.0%). "Efficacy" was ranked as the primary concern among all groups; a substantial portion of respondents reported experiencing adverse drug reactions upon generic substitution (general public: 26.6%; doctors: 23.3%; nurses: 9.1%; pharmacists: 42.0%). At least half of the general public, nurses, and pharmacists considered that patients should be given a choice for generic substitution. However, fewer than one-fifth of doctors and nurses and approximately one-third of pharmacists considered that patient consent was needed prior to generic substitution, compared with approximately two-thirds of the general public., Conclusion: The knowledge and perception of generic substitution remains low, both in the general public and among health care professionals. This knowledge deficit could potentially lead to different perspectives among stakeholders regarding generic substitution.

The group C SOX transcription factors SOX4, -11 and -12 play important and mutually overlapping roles in development of a number of organs. Here, we examined the role of SoxC genes during gonadal development in mice. All three genes were expressed in developing gonads of both sexes, predominantly in somatic cells, with Sox4 being most strongly expressed. Sox4 deficiency resulted in elongation of both ovaries and testes, and an increased number of testis cords. While female germ cells entered meiosis normally, male germ cells showed reduced levels of differentiation markers Nanos2 and Dnmt3l and increased levels of pluripotency genes Cripto and Nanog, suggesting that SOX4 may normally act to restrict the pluripotency period of male germ cells and ensure their proper differentiation. Finally, our data reveal that SOX4 (and, to a lesser extent, SOX11 and -12) repressed transcription of the sex-determining gene Sox9 via an upstream testis-specific enhancer core (TESCO) element in fetal gonads, raising the possibility that SOXC proteins may function as transcriptional repressors in a context-dependent manner., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In most eutherian mammals, sex determination is governed by the Y-linked gene Sry, but in African pygmy mice Mus minutoides, Sry action is overridden by a variant X chromosome (X*), yielding X*Y females. We hypothesized that X*Y sex reversal may be underpinned not only by neomorphic X chromosome functionality, but also by a compromised Sry pathway. Here, we show that neither M. minutoides SRY nor its target, the Sox9-TESCO enhancer, had appreciable transcriptional activity in in vitro assays, correlating with sequence degradation compared to Mus musculus counterparts. However, M. minutoides SRY activated its cognate TESCO to a moderate degree, and can clearly engage the male pathway in M. minutoides in the wild, indicating that SRY and TESCO may have co-evolved in M. minutoides to retain function above a threshold level. We suggest that weakening of the SRY/TESCO nexus may have facilitated the rise and spread of a variant X* chromosome carrying female-inducing modifier gene(s)., Competing Interests: The authors declare no competing financial interests.

Cellular enzymes interact in a post-translationally regulated fashion to govern individual cell behaviors, yet current platform technologies are limited in their ability to measure multiple enzyme activities simultaneously in single cells. Here, we developed multi-color Förster resonance energy transfer (FRET)-based enzymatic substrates and use them in a microfluidics platform to simultaneously measure multiple specific protease activities from water-in-oil droplets that contain single cells. By integrating the microfluidic platform with a computational analytical method, Proteolytic Activity Matrix Analysis (PrAMA), we are able to infer six different protease activity signals from individual cells in a high throughput manner (~100 cells/experimental run). We characterized protease activity profiles at single cell resolution for several cancer cell lines including breast cancer cell line MDA-MB-231, lung cancer cell line PC-9, and leukemia cell line K-562 using both live-cell and in-situ cell lysis assay formats, with special focus on metalloproteinases important in metastasis. The ability to measure multiple proteases secreted from or expressed in individual cells allows us to characterize cell heterogeneity and has potential applications including systems biology, pharmacology, cancer diagnosis and stem cell biology., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The technique described in this protocol allows the user to position small tissues in the optimal orientation for paraffin embedding and sectioning by first immobilising the tissue in an agarose/gelatin cube. This method is an adaptation of methods used for early embryos and can be used for any small tissues or embryo segments. Processing of larger tissue sections using molds to create agarose/gelatin blocks has been described previously; this detailed protocol provides a method for dealing with much smaller tissues or embryos (≤5mm). The tissue is briefly fixed then an agarose/gelatin drop is created to surround the tissue. The tissue can be orientated as per the user's preference in the drop before it sets as is carved into a cube with a domed top. The cube is then dehydrated and goes through the embedding and sectioning process. The domed cube is easy to orientate when embedding the tissue in a wax block giving the user assured orientation of the small tissue for sectioning. Additionally, the agarose/gelatin cube is easy to see in the unmolded wax once embedded, making the region of interest easy to identify., (Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.)

Today's ecosystems are influenced by different factors that could evolve into stressors. Effects of pesticides, especially in agricultural areas, may interact with environmental factors, such as soil moisture fluctuation caused by global climate change. In this contribution, two semi-field studies conducted in Germany and Portugal with terrestrial model ecosystems are presented. Their aim was to assess the effects of the fungicide pyrimethanil under different soil moisture levels on Enchytraeidae. In Portugal a no observed effect concentration design was chosen, using two concentration levels: the maximum application rate (MAR) according to the safe use registration within the European Union and five times the MAR (1.82 and 9.09 mg/kg dry soil, respectively). Both concentrations did neither affect the total enchytraeid abundance nor single populations. In Germany an ECx design (effect concentration) was conducted, using 11 concentrations. In general, 14 EC50 values for different combinations of single species, moisture level and sampling date were determined. The strongest effects were found in dry soil, particularly for Fridericia connata (EC50: 3.48 mg/kg dry soil after 8 weeks of exposure). The advantages and challenges of these test designs are discussed with regard to the registration process of pesticides in the European Union. In any case, enchytraeids are suitable test organisms in such higher tier studies for the combined evaluation of chemical and climatic stressors due to their usually high diversity and abundances and their close contact with the soil solution.

Background: Conservative treatment of inferior breast cancers has been a challenge for breast surgeons due to the high incidence of poor cosmetic outcomes. In 2008, Renouvel et al. described an oncoplastic 'Crescent' technique utilizing an advancement flap to fill the defect after cancer excision in the lower pole of the breast. A follow-up study demonstrated no local recurrence at 45 months and excellent or good cosmetic outcomes in over 70% of patients. This study aims to assess the outcomes of applying this 'Crescent' technique in a breast surgical unit., Methods: Retrospective study carried out at Westmead Breast Cancer Institute on 16 patients treated with the 'Crescent' technique. Data regarding patient and tumour characteristics, operative outcomes and complications were obtained. Patients were invited to complete a modified Breast-Q questionnaire and have their photographs taken to assess patient satisfaction and cosmetic outcome., Results: Over 12 months, 16 women underwent the 'Crescent' technique. Mean tumour size was 11.4 mm (range 2.0-36 mm) and median resected volume was 33.0 g (range 15-117 g). One patient (7.1%) had involved margins. One patient returned to theatre for evacuation of a haematoma. Cosmetic outcome was excellent or good in over 80% of patients., Conclusion: The oncoplastic 'Crescent' technique is a safe and reliable technique with good cosmetic outcome that can be implemented in a breast surgical unit., (© 2015 Royal Australasian College of Surgeons.)

The issues of whether and how some organs coordinate their size and shape with the blueprint of the embryo axis, while others appear to regulate their morphogenesis autonomously, remain poorly understood. Mutations in Ift144, encoding a component of the trafficking machinery of primary cilia assembly, result in a range of embryo patterning defects, affecting the limbs, skeleton and neural system. Here, we show that embryos of the mouse mutant Ift144(twt) develop gonads that are larger than wild-type. Investigation of the early patterning of the urogenital ridge revealed that the anterior-posterior domain of the gonad/mesonephros was extended at 10.5 dpc, with no change in the length of the metanephros. In XY embryos, this extension resulted in an increase in testis cord number. Moreover, we observed a concomitant extension of the trunk axis in both sexes, with no change in the length of the tail domain or somite number. Our findings support a model in which: (1) primary cilia regulate embryonic trunk elongation; (2) the length of the trunk axis determines the size of the urogenital ridges; and (3) the gonad domain is partitioned into a number of testis cords that depends on the available space, rather than being divided a predetermined number of times to generate a specific number of cords., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The mammalian sex-determining factor SRY comprises a conserved high-mobility group (HMG) box DNA-binding domain and poorly conserved regions outside the HMG box. Mouse Sry is unusual in that it includes a C-terminal polyglutamine (polyQ) tract that is absent in nonrodent SRY proteins, and yet, paradoxically, is essential for male sex determination. To dissect the molecular functions of this domain, we generated a series of Sry mutants, and studied their biochemical properties in cell lines and transgenic mouse embryos. Sry protein lacking the polyQ domain was unstable, due to proteasomal degradation. Replacing this domain with irrelevant sequences stabilized the protein but failed to restore Sry's ability to up-regulate its key target gene SRY-box 9 (Sox9) and its sex-determining function in vivo. These functions were restored only when a VP16 transactivation domain was substituted. We conclude that the polyQ domain has important roles in protein stabilization and transcriptional activation, both of which are essential for male sex determination in mice. Our data disprove the hypothesis that the conserved HMG box domain is the only functional domain of Sry, and highlight an evolutionary paradox whereby mouse Sry has evolved a novel bifunctional module to activate Sox9 directly, whereas SRY proteins in other taxa, including humans, seem to lack this ability, presumably making them dependent on partner proteins(s) to provide this function.

Melanoma metastatic to the liver has a very poor prognosis, and has traditionally been treated using systemic chemotherapy with limited efficacy. Surgery is increasingly being explored as a therapeutic option for melanoma liver metastases, with varying levels of success. A systematic review was undertaken to explore the short-term and long-term outcomes associated with hepatectomy for melanoma metastases, in addition to identifying prognostic factors favouring increased survival. All eligible studies were identified through an electronic search of Medline and Embase (January 1990-March 2013). Each study was independently analysed by two reviewers, with relevant data extracted and tabulated according to predetermined criteria. Thirteen studies were selected that fulfilled the selection criteria, with a total of 551 patients undergoing hepatic resection for melanoma metastases. Metastases to the liver occurred at a median interval of 54 months. The median perioperative morbidity and mortality were 10% (range 0-28.6%) and 0% (range 0-7.1%), respectively. The median overall survival for operative patients was 24 months, with median survival being greater in the R0 resection group (25 months; range 9.5-65.6 months) compared with the R1/2 resection group (16 months; range 11.7-29 months). Overall median 1-, 3- and 5-year survival rates were 70% (range 39-100%), 36% (range 10.2-53%) and 24% (range 3-53%), respectively. Positive prognostic factors may include single hepatic metastases, a longer time to development of hepatic metastases and R0 resection. Hepatic resection for metastatic melanoma might confer a distinct survival benefit in a select group of patients, although disease recurrence is the norm.

Lysosomes serve key degradative functions for the turnover of membrane lipids and protein components. Its biogenesis is principally dependent on exocytic traffic from the late endosome via the trans-Golgi network, and it also receives cargo to be degraded from the endocytic pathway. Membrane trafficking to the late endosome-lysosome is tightly regulated to maintain the amplitude of signalling events and cellular homeostasis. Key coordinators of lysosomal traffic include members of the Rab small GTPase family. Amongst these, Rab7, Rab9 and the more recently studied Rab22B/31 have all been reported to regulate membrane trafficking processed at the late endosome-lysosome system. We discuss what is known about the roles of these Rab proteins and their interacting partners on the regulation of traffic of important receptor proteins such as the epidermal growth factor receptor (EGFR) and the mannose 6-phosphate receptor (M6PR), in association with the late endosome-lysosome system. Better knowledge of EGFR and M6PR traffic in this regard may aid in understanding the pathological processes, such as oncogenic transformations associated with these receptors., (Copyright © 2012 John Wiley & Sons, Ltd.)

Genetic engineering has been used to modify peanut cultivars for improving agronomic performance and pest resistance. Food products developed through genetic engineering have to be assessed for their safety before approval for human consumption. Preservation of desirable chemical, flavor and aroma attributes of the peanut cultivars during the genetic modifications is critical for acceptance of genetically modified peanuts (GMP) by the food industry. Hence, the main objective of this study is to examine chemical characteristics and volatile profile of GMP. The genetically modified peanut cultivars, 188, 540 and 654 were obtained from the USDA-ARS in Stillwater, Oklahoma. The peanut variety Okrun was examined as a control. The volatile analysis was performed using a gas chromatograph/mass spectrometer (GC/MS) equipped with an olfactory detector. The peanut samples were also analyzed for their moisture, ash, protein, sugar and oil compositions. Experimental results showed that the variations in nutritional composition of peanut lines examined in this study were within the values reported for existing cultivars. There were minor differences in volatile profile among the samples. The implication of this study is significant, since it shows that peanut cultivars with greater pest and fungal resistance were successfully developed without major changes in their chemical characteristics.

The Rab family of small GTPases serves as cellular regulators of vesicular transport in all eukaryotes. Higher organisms have a vastly expanded number of Rabs. Some of these would presumably perform cell-type or tissue-specific functions. Roles for Rab proteins in several aspects of cellular physiology that are peculiar to neurons have been revealed. Rabs, in conjunction with their effectors and regulators, participate in central nervous system development (Rab23), polarized neurite growth (Rab8 and Rab11), endocytosis and axonal retrograde transport (Rab5 and Rab7). At chemical synapses, Rabs perform specific functions in synaptic vesicle exocytosis (Rab3) and postsynaptic compartment dynamics of glutamate receptors (Rab8). Much less is known about the role of Rabs in astroglia/oligodendroglia-specific processes, although Rab family members that are glia-enriched (Rab22B, Rab40C) have been reported. We summarized and discussed recent findings on the roles played by Rabs in neurons and glia in the central nervous system, and speculate on future directions.

The small GTPase Rab22B (or Rab31) has been suspected to be involved in trafficking at trans-Golgi network. However, its exact cellular localization, tissue expression profile, and functions have not been uncharacterized. Specific antibody raised against Rab22B's protein revealed that Rab22B is brain-enriched, but is also present in substantial levels in spleen and intestine. In HeLa cells, endogenous Rab22B is largely associated with the trans-Golgi network (TGN). Over-expression of a GDP-binding mutant (Rab22BSN), but not wild-type Rab22B, specifically disrupts the TGN localization of TGN46, a dynamic marker which cycles between the TGN and the plasma membrane. The TGN resident membrane protein syntaxin 16, cis-Golgi markers such as GM130 and syntaxin 5, as well as the TGN/late endosome marker mannose 6-phosphate receptor (M6PR) are not affected by Rab22BSN, neither was endosomal-TGN transport of the Shiga toxin B subunit. The disruption of TGN46 staining by Rab22BSN could be specifically attributed to a domain at the C-terminal portion of Rab22B, where its sequence deviates the most from Rab22A. Over-expression of Rab22BSN inhibits the cell surface transport of the vesicular stomatitis virus G protein. Thus, Rab22B may have a role in anterograde exit from the TGN.

Previous studies on individual differences in mathematical abilities have shown that working memory contributes to early arithmetic performance. In this study, we extended the investigation to algebraic word problem solving. A total of 151 10-year-olds were administered algebraic word problems and measures of working memory, intelligence quotient (IQ), and reading ability. Regression results were consistent with findings from the arithmetic literature showing that a literacy composite measure provided greater contribution than did executive function capacity. However, a series of path analyses showed that the overall contribution of executive function was comparable to that of literacy; the effect of executive function was mediated by that of literacy. Both the phonological loop and the visual spatial sketchpad failed to contribute directly; they contributed only indirectly by way of literacy and performance IQ, respectively.