The lived experience of young adults with Fetal Valproate Spectrum Disorder, and the perspective of their parents: A qualitative study.

BACKGROUND: While research has investigated the physical and neurodevelopmental consequences following prenatal exposure to valproate, our understanding of individuals with a formal diagnosis of Fetal Valproate Spectrum Disorder (FVSD), particularly in the context of adulthood, remains limited. AIM: To investigate how symptoms and challenges of FVSD present in adulthood. METHODS: 30 people took part in the study, including 13 young adults aged between 21 and 37 years, 15 mothers, and 2 fathers. In all cases, valproate had been used for the treatment of maternal epilepsy. Data were collected using semi-structured interviews and analysed using thematic analysis. RESULTS: Six broad themes were identified: 1. Health and development, 2. Employment, 3. Daily living and independence, 4. Social skills and relationships, 5. Access to services, and 6. Impact on families. Individuals with FVSD live with an array of physical, mental, and developmental challenges that extend well beyond childhood, significantly altering their life course and that of their families. Challenges in obtaining employment, achieving independent living, and navigating social and romantic relationships become increasingly significant as individuals with FVSD age. Despite their persistent need for support, services for adults with FVSD are either limited or entirely absent. Recommendations from families were provided regarding optimized support systems. CONCLUSION: This study highlights the lifelong physical, cognitive, emotional, social and behavioural symptoms associated with FVSD. Young adults and their parents desire further research regarding the condition along with improved support and health services in adulthood.

Does gene expression in laryngeal subsites differ between patients with laryngopharyngeal reflux and controls?

OBJECTIVE: To identify laryngeal mRNA gene changes in patients with laryngopharyngeal reflux (LPR). METHOD: Laryngeal biopsies from non-smoking LPR patients (n=10; Reflux Symptom Index (RSI) >12 and a Reflux Finding Score (RFS) >6) and controls (n=9; RSI <12 and RFS <6) were collected from four subsites (true vocal cord, false vocal cord, medial arytenoid and posterior commissure) of the larynx. qRT-PCR analyses were conducted on 20 reflux- and inflammation-related genes, including interleukins 6 and 8, cytokeratins 8 and 14, mucin genes MUC1, MUC2, MUC3B, MUC4, MUC5B, MUC6 and MUC7 and carbonic anhydrase III. Statistical analysis (Mann-Whitney U test) compared gene expression levels between LPR and control groups at each subsite. RESULTS: Site-specific differences in squamous metaplasia and gene expression were noted in LPR patients, with the majority present in the medial arytenoid region. Significant.differences were noted in genes related to mucosal defence and inflammation, including CRNN, CD1d, TGFß-1, MUC2, MUC5B and CDH1. CONCLUSION: Whilst the posterior commissure is commonly identified as the area demonstrating the most significant macroscopic change in LPR, the histological changes and genes assessed here showed more pronounced LPR associated differences in the medial arytenoid. We identified differences in expression of mucin genes, cytokeratin-14 and molecular markers of inflammation. Whilst some of these changes may be metaplasia-related, further evaluation of the mRNA expression of these genes may provide a useful biomarker panel for diagnosis and therapeutic monitoring of LPR.

Neurodevelopmental outcomes in children and adults with Fetal Valproate Spectrum Disorder: A contribution from the ConcePTION project.

AIM: To describe the neurodevelopmental phenotype of older children and adults with a diagnosis of Fetal Valproate Spectrum Disorder (FVSD). METHODS: In this cross-sectional study, 90 caregivers were recruited and completed a series of questionnaires regarding the neurodevelopmental outcomes of 146 individuals aged 7-37 years (M = 18.1), including individuals with a formal diagnosis of FVSD (n = 99), individuals exposed to Valproate but without an FVSD diagnosis (n = 24), and individuals not exposed to Valproate (N = 23). The mean dose of valproate exposure for individuals with an FVSD diagnosis was 1470 mg/day. RESULTS: Individuals with a diagnosis of FVSD showed significantly higher levels of moderate (43.4%) and severe (14.4%) cognitive impairment than other groups (p = 0.003), high levels of required formal educational support (77.6%), and poorer academic competence than individuals not exposed to Valproate (p = 0.001). Overall psychosocial problems (p = 0.02), internalising problems (p = 0.05) and attention problems (p = 0.001), but not externalising problems, were elevated in individuals with a diagnosis of FVSD. Rates of neurodevelopmental disorders, particularly autistic spectrum disorders (62.9%) and sensory problems (80.6%) are particularly central to the FVSD phenotype. There was no evidence of a statistical dose-dependent effect, possibly due to the high mean dose of exposure having a uniformly negative impact across the sample. Individuals with FVSD had required a significant number of health and child development services. INTERPRETATION: Children and young adults with a diagnosis of FVSD are at an increased risk of a range of altered neurodevelopmental outcomes, highlighting the need for a multidisciplinary approach to clinical management across the lifespan.

Tamoxifen enhances the cytotoxicity of conventional chemotherapy in esophageal adenocarcinoma cells.

INTRODUCTION: Esophageal adenocarcinoma is a lethal malignancy which is increasing in incidence, and many patients receive chemotherapy as part of their treatment. We have previously demonstrated that esophageal adenocarcinoma-derived cell lines respond to treatment with estrogen receptor modulators, such as tamoxifen. Reports from breast cancer suggest that tamoxifen may attenuate the efficacy of other chemotherapeutic agents. We have therefore assessed the response of esophageal adenocarcinoma cell lines to tamoxifen therapy when given in combination with conventional agents. METHODS: Two estrogen receptor (ER)-positive esophageal adenocarcinoma cell lines (OE-19 and OE-33) were treated with combinations of tamoxifen, cisplatin and 5-fluorouracil (5-FU). Effects on cell viability were measured using an MTS assay, and cell death was detected with annexin V/propidium iodide flow cytometry. To assess whether the efficacy of tamoxifen in these cell lines might be relevant to the clinical setting, we analyzed ER status in 10 esophageal adenocarcinoma tissue specimens by immunohistochemistry. RESULTS: IC50 values (µM) for OE-19 and OE-33 were 11.2 and 7.1 for tamoxifen, 19.6 and 4.7 for cisplatin, and 1.7 and 5.9 for 5-FU, respectively. Cell death was detected in 11.9% and 15.8% of cells treated with tamoxifen, 7.9% and 8.7% cells treated with cisplatin, and 3.6% and 8.6% cells treated with 5-FU at their IC50s. The addition of tamoxifen to cisplatin increased cell death by 11.4% in OE-19 (p < 0.0001) and 16.3% in OE-33 (p < 0.0001). Similarly, the addition of tamoxifen to 5-FU increased cell death by 11.6% in OE-19 (p < 0.0001) and 15.9% in OE-33 (p < 0.0001). Eight of 10 tissue specimens showed positive staining for ERα and 7 of 10 for ERß. CONCLUSIONS: In a cell culture model the addition of tamoxifen to conventional chemotherapy appears to be both feasible and beneficial. Expression of ERα and ERß was also confirmed in esophageal adenocarcinoma tissues.

Characteristics of skeletal muscle chloride channel C1C-1 and point mutant R304E expressed in Sf-9 insect cells.

Using the baculovirus system, the skeletal muscle chloride channel, CIC-1 (rat), and a point mutant replacing arginine 304 with glutamic acid were expressed at high levels in cultured Sf-9 insect cells. Whole-cell patch-clamping revealed large inwardly rectifying currents with maxima up to 15 nA inward and 2.5 nA outward. Saturation was evident at voltage steps positive to +40 mV whilst steps negative to -60 mV produced inactivating currents made up of a steady state component and two exponentially decaying components with tau 1 = 6.14+/- 0.92 ms, tau 2 = 36.5+/- 3.29 ms (S.D) n = 7 for steps to -120 mV. Currents recorded in the outside-out patch configuration were often unexpectedly large and up to 5% of whole-cell currents obtained in the same cell, suggesting an uneven channel distribution in the plasmalemma of Sf-9 cells. The pharmacology of a number of chloride channel blockers, including anthracene-9-carboxylate (A9C), niflumate, and perrhenate, was investigated and showed for the first time that perrhenate is an effective blocker of C1C-1 and that it has a complex mechanism of action. Further, the potency of A9C was found to be dependent on external chloride concentration. As in studies on muscle cells themselves, blockade was rapidly effective and easily reversible, except when applying the indanyloxyacetate derivative, IAA94/95, which took up to 10 min to act, and, consistent with an intracellular site of action, was difficult to reverse by washing. Mutation of the highly conserved arginine at position 304 to a glutamic acid did not significantly alter the behaviour of the channel.

Modulation of the gating of CIC-1 by S-(-) 2-(4-chlorophenoxy) propionic acid.

1. Using whole-cell patch-clamping and Sf-9 cells expressing the rat skeletal muscle chloride channel, rCIC-1, the cellular mechanism responsible for the myotonic side effects of clofibrate derivatives was examined. 2. RS-(+/-) 2-(4-chlorophenoxy)propionic acid (RS-(+/-) CPP) and its S-(-) enantiomer produced pronounced effects on CIC-1 gating. Both compounds caused the channels to deactivate more rapidly at hyperpolarizing potentials, which showed as a decrease in the time constants of both the fast and slow deactivating components of the whole cell currents. Both compounds also produced a concentration-dependent shift in the voltage dependence of channel apparent open probability to more depolarizing potentials, with an EC50 of 0.79 and 0.21 mM for the racemate and S-(-) enantiomer respectively. R-(+) CPP at similar concentrations had no effect on gating. RS-(+/-) CPP did not block the passage of Cl- through the pore of rCIC-1. 3. CIC-1 is gated by Cl- binding to a site within an access channel and S-(-) CPP alters gating of the channel by decreasing the affinity of this binding site for Cl-. Comparison of the EC50 for RS-(+/-) CPP and S-(-) CPP indicates that R-(+) CPP can compete with the S-(-) enantiomer for the site but that it is without biological activity. 4. RS-(+/-) CPP produced the same effect on rCIC-1 gating when added to the interior of the cell and in the extracellular solution. 5. S-(-) CPP modulates the gating of CIC-1 to decrease the membrane Cl- conductance (GCl), which would account for the myotonic side effects of clofibrate and its derivatives.

Characterization of the small cryptic plasmid, pIMVS1, of Salmonella enterica ser. Typhimurium.

The nucleotide sequence has been determined of a small cryptic plasmid of Salmonella Typhimurium, designated pIMVS1, which correlated with an outbreak of gastroenteritis. This plasmid has a size of 3357 bp with no known functions. It does not encode any protein products but shows homology to several well-characterized plasmids. The replication system with RNAI and RNAII and the replication origin (oriV) is highly similar to that of plasmid p15A. It also has a putative mobilisation origin similar to that of ColE1.

Comparison of improved BACTEC and Lowenstein-Jensen media for culture of mycobacteria from clinical specimens.

A 4-month trial involving 2,563 routine clinical specimens was conducted to compare the improved BACTEC TB system (12B medium) with the conventional Lowenstein-Jensen (LJ) media for the isolation, identification, and susceptibility testing of mycobacteria. One hundred sixty-two mycobacterial isolates were recovered, 147 (91%) with BACTEC and 118 (73%) with LJ media. Of these, 62 were Mycobacterium tuberculosis complex strains, 59 (95%) of which were isolated with BACTEC and 54 (87%) of which were isolated with LJ media. Of the remaining 100 isolates, which were mycobacteria other than tuberculosis (MOTT), BACTEC and LJ media detected 88 and 64%, respectively. The contamination rate was significantly higher in BACTEC (5%) than in LJ media (3.3%). The mean isolation time for M. tuberculosis complex with BACTEC was 15.5 days, compared with 25.6 days with LJ. For MOTT, the mean isolation times were 5.8 and 21.4 days, respectively. Identification of 32 M. tuberculosis complex isolates and 38 isolates of MOTT by the BACTEC NAP (p-nitro-alpha-acetylamino-beta-hydroxypropiophenone) inhibition test gave 100% agreement with conventional biochemical identifications. The results of susceptibility testing of 18 M. tuberculosis complex isolates with BACTEC agreed completely with those obtained by the resistance ratio method.

pH-dependent interactions of Cd2+ and a carboxylate blocker with the rat C1C-1 chloride channel and its R304E mutant in the Sf-9 insect cell line.

1. Gating of the skeletal muscle chloride channel (ClC-1) is sensitive to extracellular pH. In this study, whole-cell recording of currents from wild-type (WT) ClC-1 and a mutant, R304E, expressed in the Sf-9 insect cell line was used to investigate further the nature of the pH-sensitive residues. 2. Extracellular Cd2+ produced a concentration-dependent block of WT ClC-1 with an IC50 of 1.0 +/- 0.1 mM and a Hill coefficient of 2.0 +/- 0.3. This block was sensitive to external pH, reducing at low pH, with an apparent pKa of 6.8 +/- 0.1 and a Hill coefficient for proton binding of 3.0 +/- 0.3. Anthracene-9-carboxylate (A-9-C) block of WT ClC-1 was also pH sensitive, increasing at low pH, with an apparent pKa of 6.4 +/- 0.1 and a Hill coefficient for proton binding of 1.0 +/- 0.2. 3. Compared with WT ClC-1, R304E had a lower affinity for Cd2+ (IC50, 3.0 +/- 0.3 mM) but it had a similar Hill coefficient for transition metal ion binding. The Hill coefficient for proton binding to the Cd2+ binding site was reduced to 1.4 +/- 0.3. In contrast, the A-9-C binding site in R304E showed the same pH sensitivity and affinity for the blocker as that seen in WT ClC-1. 4. ClC-1 has at least two binding sites for Cd2+, each of which has at least three residues which can be protonated. Binding of A-9-C is influenced by protonation of a single residue. Arg 304 is not sufficiently close to the A-9-C binding site to affect its characteristics, but it does. alter Cd2+ binding, indicating that transition metal ions and aromatic carboxylates interact with distinct sites. 5. The block of ClC-1 by transition metal ions and the apparent pKa of this block, together with the apparent pKa for A-9-C block and gating are all compatible with the involvement of His residues in the pore and gate of ClC-1.

Concentration and pH dependence of skeletal muscle chloride channel ClC-1.

1. The influence of Cl- concentration and pH on gating of the skeletal muscle Cl- channel, ClC-1, has been assessed using the voltage-clamp technique and the Sf-9 insect cell and Xenopus oocyte expression systems. 2. Hyperpolarization induces deactivating inward currents comprising a steady-state component and two exponentially decaying components, of which the faster is weakly voltage dependent and the slower strongly voltage dependent. 3. Open probability (Po) and kinetics depend on external but not internal Cl- concentration. 4. A point mutation, K585E, in human ClC-1, equivalent to a previously described mutation in the Torpedo electroplaque chloride channel, ClC-0, alters the I-V relationship and kinetics, but retains external Cl- dependence. 5. When external pH is reduced, the deactivating inward currents of ClC-1 are diminished without change in time constants while the steady-state component is enhanced. 6. In contrast, reduced internal pH slows deactivating current kinetics as its most immediately obvious action and the Po curve is shifted in the hyperpolarizing direction. Addition of internal benzoate at low internal pH counteracts both these effects. 7. A current activated by hyperpolarization can be revealed at an external pH of 5.5 in ClC-1, which in some ways resembles currents due to the slow gates of ClC-0. 8. Gating appears to be controlled by a Cl(-)-binding site accessible only from the exterior and, possibly, by modification of this site by external protonation. Intracellular hydroxyl ions strongly affect gating either allosterically or by direct binding and blocking of the pore, an action mimicked by intracellular benzoate.