Additional file 2 of Exploring culinary medicine as a promising method of nutritional education in medical school: a scoping review

Additional file 2

Additional file 1 of Exploring culinary medicine as a promising method of nutritional education in medical school: a scoping review

Additional file 1

Molecular and cellular defects in <i>Sr/+;Nr/+</i> SCO development at 12.5 dpc and 14.5 dpc.

<p><b>A,B:</b> Saggital section of 12.5 dpc wild type (A) and <i>Sr/+;Nr/+</i> (B) SCO region showing immuno-positive BrdU cells. <b>C:</b> Quantitation of BrdU showing a significant increase in proliferation in the <i>Sr/+;Nr/+</i> SCO. <b>D–F:</b> Section <i>in situ</i> hybridisation analysis of <i>Fibcd1</i> expression in 12.5 dpc (D), 15.5 dpc (E) and 18.5 dpc (F) SCO. <b>G,H:</b> Sagittal section <i>in situ</i> hybridisation analysis showing decreased <i>Fibcd1</i> expression in the <i>Sr/+;Nr/+</i> (H) SCO compared with wild type SCO (G) at 12.5 dpc. <b>I,J:</b> In situ hybridisation analysis of wild type (I) and <i>Sr/+;Nr/+</i> (J) 14.5 dpc sagittal sections showing robust <i>Fibcd1</i> expression in the wild type SCO but not in the dysplasic <i>Sr/+;Nr/+</i> SCO region (hash). A small group of <i>Fibcd1</i> positive cells were present at the posterior limit of the <i>Sr/+;Nr/+</i> SCO region (indicated by the plus sign). <b>K:</b> qRT-PCR analysis of SCO marker expression in 12.5 dpc wild type and <i>Sr/+;Nr/+</i> SCO biopsies. <b>L–O:</b> Whole mount <i>in situ</i> hybridisation showing <i>Msx1</i> (L, N) and <i>Wnt1</i> (M,O) expression at the dorsal midline of wild type (L,M) and <i>Sr/+;Nr/+</i> (N,O) embryos at 12.5 dpc. <i>Wnt1</i> and <i>Msx1</i> expression was lower in <i>Sr/+;Nr/+</i> embryos rostral to the posterior limit of SCO (arrowhead). <b>P, Q: </b><i>In situ</i> hybridisation analysis of coronal sections from 12.5 dpc wild type (P) and <i>Sr/+;Nr/+</i> (Q) embryos showing dramatic downregulation of <i>Bmp6</i> expression in the transgenic SCO. <b>R–U: </b><i>In situ</i> hybridisation of coronal sections from 12.5 dpc wild type (R and T) and <i>Sr/+;Nr/+</i> (S and U) demonstrating the dorsal domain of dorsolateral neuroepithelial cells has expanded ventrally. MHN: medial habenular nucleus. PC: posterior commissure. Arrows point to the SCO primordium invagination. Double-headed arrows indicate the mesocoelic recess. In A, B, D–J and L–O, anterior is to the left and dorsal to the top. Scale bars are 100 µm (A–B, D–H), 200 µm (I–J, P–U) and 500 µm (L–O).</p

Defective SCO development in <i>Sr/+;Nr/+</i> embryos from 13.5 dpc.

<p><b>A–D:</b> Nissl stained sagittal sections of 12.5 dpc wild type (A,C) and <i>Sr/+;Nr/+</i> (B,D) SCO primordia showing similar morphology and cell body density. C and D are boxed regions in A and B, respectively. <b>E–H:</b> Nissl-stained sagittal sections of 13.5 dpc wild type (E,G) and <i>Sr/+;Nr/+</i> (F,H) SCO region. Note the thinner SCO primordium and lack of pineal recess in the <i>Sr/+;Nr/+</i> embryo. <b>I–N:</b> Nissl-stained sagittal (I,J) and coronal (K–N) sections of 15.5 dpc wild type (I,K,M) and <i>Sr/+;Nr/+</i> (J,L,N) SCO region. Note the severely disrupted PC and absence of pseudostratified ependymal layer of the SCO in the <i>Sr/+;Nr/+</i> embryo. M and N are boxed regions in K and L, respectively. <b>O–P:</b> RF (red) and SOX3 (green) immunostaining of 15.5 dpc sagittal wild type (O) and <i>Sr/+;Nr/+</i> (P) sections. <i>Sr/+;Nr/+</i> transgenic embryos maintain SOX3 expression in the dorsal midline but fail to generate RF. Arrows point to the SCO invagination in the roof plate and the double-headed arrow to the mesocoelic recess. The hash sign indicates the SCO remnant. The asterisk indicates the approximate position where the pineal gland should develop in <i>Sr/+;Nr/+</i> embryos. The arrowhead indicates the pineal recess. EpTh: epithalamus. MHN: medial habenular nucleus. Pt: pretectum. FR: fasciculus retroflexus. PC: posterior commissure. In A–H and K–P, anterior is to the left and dorsal to the top. Scale bars are 100 µm (C–F, M,N), 200 µm (A,B,G,H,K,L) and 1 mm (I,J).</p

Structure, copy number and expression of the <i>Sox3</i> transgene.

<p><b>A:</b> Schematic representation of the transgene showing the <i>Sox3</i> coding sequence (which is contained in a single exon) and the <i>IRES</i>-<i>EGFP</i> reporter cassette and flanking genomic sequence (not to scale). IRES indicates the internal ribosome entry site. <b>B:</b> qPCR analysis of transgene copy number in the <i>Sr/+</i> and <i>Nr/+</i> lines demonstrating that they contain 2 and 1 copy of the transgene, respectively. <i>Sox3</i> null (KO) genomic DNA was used as a negative control <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029041#pone.0029041-Rizzoti1" target="_blank">[22]</a>. <i>Sox1</i> quantification was performed as an endogenous autosomal control. <b>C:</b> Quantitative fluorescence Western blot analysis showing a 2–3-fold increase in SOX3 protein levels in 10.5 dpc <i>Sr/+</i> and <i>Nr/+</i> embryos.</p

Dose-dependent penetrance of overt CH in <i>Sox3</i> transgenic mice.

<p>• and ∧: Significantly fewer progeny were present at weaning (<i>*p</i> = 0.030, ∧<i>p</i> = 0.001; Chi Square test).</p><p>Table showing the penetrance of overt CH in <i>Sr/</i>, <i>Nr/+</i> and <i>Sr/+;Nr/+</i> pups at weaning (approximately 3 weeks).</p

<i>Sox3</i> transgenic mice displayed CH.

<p><b>A,B:</b> H&E stained coronal section of adult <i>Sox3</i> transgenic mouse brain without (A) and with (B) overt hydrocephalus. Note the expanded lateral ventricles (LV), thinning of cerebral cortex (double-headed arrow) and hippocampal deformation (arrows) in the CH founder. <b>C,D: </b><i>Nr/+</i> adult mice showing dome-shaped cranium (arrow) due to overt CH (D) and normal cranial morphology (C). <b>E,F:</b> Nissl stained 18.5 dpc coronal brain sections of wild type (E) and <i>Sr/+;Nr/+</i> (F) 18.5 dpc embryonic brains. Note the expansion of the lateral ventricles (LV) in the transgenic embryo (F). Scale bar: 2 mm (A–B) and 1 mm (E–F).</p

Endogenous <i>Sox3</i> and the <i>Sox3</i> transgene are expressed in the developing and mature SCO.

<p><b>A–H:</b> Sagittal sections of 12.5 dpc (A–D) and 15.5 dpc (E–H) wild type (A,C,E,G) and <i>Nr</i>/+ (B,D,F,H) embryos immunostained for SOX3 (A,B,E,F) and EGFP (C,D,G,H). SOX3 is expressed in all cells of the wild type and <i>Nr/+</i> SCO. Note the presence of the EGFP signal and higher intensity of SOX3 signal in Nr/+ transgenic sections. <b>I–L:</b> Mid-sagittal sections of adult wild type (I,K) and <i>Nr</i>/+ (J,L) brains showing expression of SOX3 and EGFP. <b>M–P:</b> Coronal sections of 12.5 dpc wild type (M,O) and <i>Sr/+;Nr/+</i> transgenic (N,P) SCO tissue. In wild type embryos, a lower level of endogenous SOX3 was detected in the SCO primordium in comparison with periluminal cells flanking the SCO (asterisk in M). This difference in SOX3 signal intensity was not observed in <i>Sr/+;Nr/+</i> embryos (N). Arrows: PC (posterior commissure). Yellow arrowheads: points of invagination of the SCO primordium. White arrowhead: mesocoelic recess. In A–L, anterior is to the left and dorsal to the top. Scale bar: 200 µm.</p