Z-disc protein CHAPb induces cardiomyopathy and contractile dysfunction in the postnatal heart

The Z-disc is a crucial structure of the sarcomere and is implicated in mechanosensation/transduction. Dysregulation of Z-disc proteins often result in cardiomyopathy. We have previously shown that the Z-disc protein Cytoskeletal Heart-enriched Actin-associated Protein (CHAP) is essential for cardiac and skeletal muscle development. Furthermore, the CHAP gene has been associated with atrial fibrillation in humans. Here, we studied the misregulated expression of CHAP isoforms in heart disease. Mice that underwent transverse aortic constriction and calcineurin transgenic (Tg) mice, both models of experimental heart failure, displayed a significant increase in cardiac expression of fetal isoform CHAPb. To investigate whether increased expression of CHAPb postnatally is sufficient to induce cardiomyopathy, we generated CHAPb Tg mice under the control of the cardiac-specific αMHC promoter. CHAPb Tg mice displayed cardiac hypertrophy, interstitial fibrosis and enlargement of the left atrium at three months, which was more pronounced at the age of six months. Hypertrophy and fibrosis were confirmed by evidence of activation of the hypertrophic gene program (Nppa, Nppb, Myh7) and increased collagen expression, respectively. Connexin40 and 43 were downregulated in the left atrium, which was associated with delayed atrioventricular conduction. Tg hearts displayed both systolic and diastolic dysfunction partly caused by impaired sarcomere function evident from a reduced force generating capacity of single cardiomyocytes. This co-incided with activation of the actin signalling pathway leading to the formation of stress fibers. This study demonstrated that the fetal isoform CHAPb initiates progression towards cardiac hypertrophy, which is accompanied by delayed atrioventricular conduction and diastolic dysfunction. Moreover, CHAP may be a novel therapeutic target or candidate gene for screening in cardiomyopathies and atrial fibrillatio

Long-term neurodevelopment in children with resected congenital lung abnormalities

To determine whether children who underwent resection of a congenital lung abnormality (CLA) are at higher risk for neurodevelopmental impairments than peers in the general population. The study population consisted of children born between 1999–2018 who underwent resection of a symptomatic CLA. Neurocognitive development (intelligence, memory, attention, visuospatial processing, executive functioning) and motor function of this population are monitored through our structured, prospective longitudinal follow-up program at the ages of 30 months, 5, 8, and 12 years. We compared study population scores with Dutch norm values using one-sample t-tests and one-sample binominal proportion tests. Forty-seven children were analyzed. The 8-year-olds showed significant impairments in sustained attention through the Dot Cancellation Test (mean z-scores -2.4; [-4.1; -0.8], p = 0.006 and -7.1; [-12.8; -1.4], p = 0.02 for execution speed and fluctuations respectively). Visuospatial memory was impaired at 8 years, though only in 1 out of 3 assessment tools (Rey Complex Figure Test z-scores (-1.0; [-1.5; -0.5], p < 0.001). Further neurocognitive outcomes were unimpaired at all tested ages. Regarding motor function outcomes, mean z-scores of total motor functioning were unimpaired across assessed ages. However, at 8 years, significantly more children than expected had definite motor problems (18% vs 5%, 95% CI [0.052; 0.403], p = 0.022). Conclusion: This evaluation reveals impairment in some subtests of sustained attention, visuospatial memory and motor development. However, globally, normal neurodevelopmental outcomes were found throughout childhood. We recommend testing for neurodevelopmental impairments in children who underwent surgery for CLA only if associated morbidities are present or if caregivers express doubts about their daily functioning. What is Known: • In general, surgically managed CLA cases seldom suffer from long-term surgery-related morbidity and show favorable lung function. What is New: • Long-term neurocognitive and motor function outcome appear unimpaired within surgically managed CLA cases. We recommend testing for neurodevelopmental impairments in children who underwent surgery for CLA only if associated morbidities are present or if caregivers express doubts about their daily functioning

Livelihood trade-offs in the commercialisation of multiple-use NTFP: lessons from marula (Sclerocarya birrea subsp. caffra) in southern Africa

Commercialisation of non-timber forest products (NTFP), apart from the multitude of benefits, is often associated with trade-offs in terms of traditional and cultural livelihoods. This paper presents a holistic assessment of livelihood trade-offs involved in commercialisation of marula (Sclerocarya birrea subsp. caffra), a multiple-use NTFP species in southern Africa. The study was conducted\ud at two sites in South Africa (Bushbuckridge district and Ubombo district) and one in Namibia (former Ovamboland). Some of the key features of the study include the household use and trade in marula products, the biological aspects of the marula resources, marketing and trade of the species and policies associated with its utilisation. The paper also highlights the important and diverse role that marula\ud has in local livelihoods and in contributing to the forms of livelihood capital like human, social, financial, natural and physical capital. Likely trade-offs in terms of\ud livelihoods with increasing commercialisation of marula are discussed, along with potential threats and opportunities from commercialisation

Persisting Motor Function Problems in School-Aged Survivors of Congenital Diaphragmatic Hernia

Background and Objectives: Children born with congenital diaphragmatic hernia (CDH) and treated with extracorporeal membrane oxygenation (ECMO), are at risk for motor function impairment during childhood. We hypothesized that all children born with CDH are at risk for persistent motor function impairment, irrespective of ECMO-treatment. We longitudinally assessed these children's motor function. Methods: Children with CDH with and without ECMO-treatment, born 1999–2007, who joined our structural prospective follow-up program were assessed with the Movement Assessment Battery for Children (M-ABC) at 5, 8, 12 years. Z-scores were used in a general linear model for longitudinal analysis. Results: We included 55 children, of whom 25 had been treated with ECMO. Forty-three (78%) were evaluated at three ages. Estimated mean (95% CI) z-scores from the general linear model were −0.67 (−0.96 to −0.39) at 5 years of age, −0.35 (−0.65 to −0.05) at 8 years, and −0.46 (−0.76 to −0.17) at 12 years. The 5- and 8-years scores differed significantly (p = 0.02). Motor development was significantly below the norm in non-ECMO treated patients at five years; −0.44 (−0.83 to −0.05), and at all ages in the ECMO-treated-patients: −0.90 (−1.32 to −0.49), −0.45 (−0.90 to −0.02) and −0.75 (−1.2 to −0.34) at 5, 8, and 12 years, respectively. Length of hospital stay was negatively associated with estimated total z-score M-ABC (p = 0.004 multivariate analysis). Conclusion: School-age children born with CDH are at risk for motor function impairment, which persists in those who received ECMO-treatment. Especially for them long-term follow up is recommended

Livelihood trade-offs in the commercialisation of multiple-use NTFP: lessons from marula (Sclerocarya birrea subsp. caffra) in southern Africa

Commercialisation of non-timber forest products (NTFP), apart from the multitude of benefits, is often associated with trade-offs in terms of traditional and cultural livelihoods. This paper presents a holistic assessment of livelihood trade-offs involved in commercialisation of marula (Sclerocarya birrea subsp. caffra), a multiple-use NTFP species in southern Africa. The study was conducted at two sites in South Africa (Bushbuckridge district and Ubombo district) and one in Namibia (former Ovamboland). Some of the key features of the study include the household use and trade in marula products, the biological aspects of the marula resources, marketing and trade of the species and policies associated with its utilisation. The paper also highlights the important and diverse role that marula has in local livelihoods and in contributing to the forms of livelihood capital like human, social, financial, natural and physical capital. Likely trade-offs in terms of livelihoods with increasing commercialisation of marula are discussed, along with potential threats and opportunities from commercialisation

CHAPb Tg mice show decrease cardiac performance.

<p><b>(</b>A) Representative 4-chamber view MRI images of wt and CHAPb Tg heart. Enlarged LA in CHAPb Tg is indicated with *. (B-G) MRI measurements of the left ventricle of wt (n = 4, white bars) and CHAPb Tg (n = 5, black bars) animals at 6 months of age. ED volume (B), ES volume (C), ejection fraction (D), cardiac output (E), LV mass ED (F) and LV mass ES (G). (H) Experimental setup for single membrane-permeabilized cardiomyocyte measurements to determine sarcomere function. Sarcomere force measurements at one month of age showed no difference in passive force (I; Fpas), while maximum force (J; Fmax) and Ca²⁺-sensitivity (K; pCa₅₀) were reduced in CHAPb Tg (n = 11, black bars) compared to wt (n = 12, white bars). ED (end diastolic), ES (end systolic) and LV (left ventricular). <i>t</i>-test: *, p<0.05; **, p<0.01; ***, p<0.001.</p

Sarcomeric organization is disturbed and actin signalling is increased in CHAPb Tg hearts.

<p>(A) Immunostaining for CHAP (green) and α-actinin (red) show stress fiber formation (white arrows) in CHAP Tg hearts (B) Electron microscopy analysis of wt and CHAPb Tg hearts at 6 months of age. In CHAPb Tg the sarcomeres were irregular and Z-discs and intercalated discs (black arrow heads) are disorganized, while M-bands are absent (white arrow heads) (C) Wt and CHAPb Tg hearts at 6 months of age stained for RhoA (red). In wt mice RhoA is localized at the membrane of cardiomyocytes and shows a sarcomeric expression pattern. In CHAPb Tg hearts sarcomeric expression of RhoA is absent and membrane expression is increased. (D) Western blot analysis of 2 wt and 3 CHAPb Tg hearts at 6 months of age for RhoA (24kDa), α-actinin (100kDa), actin (42kDa), Ezrin(80 kDa)/moesin(80 kDa)/radixin (75 kDa; ERM), cofilin (19kDa), SRF (40 – 67kDa) and MEF2 (40-65kDa). GAPDH (38kDa) is used as loading control. (E) Working model: in adult wt mice CHAPa is localized at the Z-disc, leading to abundance of monomeric G-actin and subsequent low expression of SRF target genes. In CHAPb Tg mice CHAPb expression results in activation of RhoA, leading to a shift from G-actin to F-actin, binding of co-factors to SRF and activation of SRF target genes, such as ANF, BNP and β-MHC. Scale bars 20 μm in A and C, 1 μm in B.</p

Chap is upregulated in mouse models of pathological cardiac hypertrophy.

<p>(A) Quantitative RT-PCR showing increased mRNA levels of both ChapA and B in hearts of transgenic mice expressing constitutively active calcineurin A (CnA Tg) (WT: n = 5, CnA Tg: n = 6; Students <i>t-test</i>: CnA Tg vs WT). (B) Western blot for CHAPa and CHAPb in adult wildtype hearts CnA Tg hearts (WT: n = 5, CnA Tg: n = 6). In wildtype hearts, CHAPa is the dominant isoform while in CnA Tg hearts both CHAPa and b were significantly upregulated. (C) Quantitative RT-PCR for ChapA showing mice subjected to transverse aortic constriction (TAC) for 1 day, 1 week and 3 weeks (n = 4, n = 4, and n = 10 respectively), with no significant regulation of ChapA mRNA compared to sham controls (n = 3, n = 3 and n = 7 respectively). (D) Quantitative RT-PCR for ChapB showing mice subjected to TAC with significant upregulation of ChapB mRNA (number of mice are same as in C). Gapdh was used as internal control in all experiments. <i>T-test</i> with Welch’s correction TAC vs sham: *, p<0.05; **, p<0.01;***, p<0.001.</p

Hypertrophy and left atrial enlargement in CHAP Tg hearts.

<p>(A-C) Wt (left panels) and CHAPb Tg (right panels) at 3 and 6 months of age. (A) HE stained overview section. The left atrium in CHAPb Tg hearts is enlarged (indicated by *) compared to wt litter mates. (B) Higher magnification of left ventricle. In the left ventricle of CHAPb Tg hearts the cardiomyocytes are hypertrophic. (C) Sirius red staining of the left ventricle showing increase in interstitial fibrosis in CHAPb Tg. (D) Myocardial volume of the left atrium (left panels) and right atrium (right panels) in wt (white bars) and CHAPb Tg (black bars) hearts at 3 months of age (wt n = 4, Tg n = 4, T-test p< 0.001). (E) CHAPb Tg heart with severe phenotype showing pronounced atrial enlargement, filled by a thrombus and thickening of the ventricles. Scale bars 1 mm in A and D, 50 μm in B, C.</p