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Histopathology reveals correlative and unique phenotypes in a high-throughput mouse phenotyping screen.

Adissu HA, Estabel J, Sunter D, Tuck E, Hooks Y, Carragher DM, Clarke K, Karp NA, Sanger Mouse Genetics ProjectNewbigging S, Jones N, Morikawa L, White JK, McKerlie C - Dis Model Mech (2014)

Bottom Line: In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen.Three of these seven lines had no clinical phenotype detected by the standard primary screen.Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls.

View Article: PubMed Central - PubMed

Affiliation: Centre for Modeling Human Disease, Toronto Centre for Phenogenomics, 25 Orde Street, Toronto, ON M5T 3H7, Canada.

ABSTRACT
The Mouse Genetics Project (MGP) at the Wellcome Trust Sanger Institute aims to generate and phenotype over 800 genetically modified mouse lines over the next 5 years to gain a better understanding of mammalian gene function and provide an invaluable resource to the scientific community for follow-up studies. Phenotyping includes the generation of a standardized biobank of paraffin-embedded tissues for each mouse line, but histopathology is not routinely performed. In collaboration with the Pathology Core of the Centre for Modeling Human Disease (CMHD) we report the utility of histopathology in a high-throughput primary phenotyping screen. Histopathology was assessed in an unbiased selection of 50 mouse lines with (n=30) or without (n=20) clinical phenotypes detected by the standard MGP primary phenotyping screen. Our findings revealed that histopathology added correlating morphological data in 19 of 30 lines (63.3%) in which the primary screen detected a phenotype. In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen. Three of these seven lines had no clinical phenotype detected by the standard primary screen. Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls. These findings demonstrate the complementary and unique contribution of histopathology to high-throughput primary phenotyping of mutant mice.

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Histopathology complements clinical phenotypes from the primary screen.Lrig−/− mice were annotated to have abnormal skin (data from male mice is shown) (Fisher’s exact test adjusted P-value=2.09e–10). The number and proportion of affected mice in knockout and wild-type baseline controls is presented (A). Abnormal skin with scaly foci (arrows) was observed in Lrig−/− mice (B). Histopathology revealed epidermal hyperplasia (arrow) and hyperkeratosis (arrowhead) (C). Mcph1−/− mice were annotated as infertile. Ovarian hypoplasia with absence of folliculogenesis was observed in Mcph1−/− females (D, top right panel); ovary from a wild-type mouse with growing follicles (arrows) is shown (D, top left panel). Seminiferous tubule vacuolation with lack of germ cells (arrows) was observed in Mcph1−/− males (D, bottom right panel); normal testis with abundant developing germ cells (arrows) is shown from a wild-type mouse (D, bottom left panel). Tbc1d10a−/− mice were annotated to have decreased circulating low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol (LDL data from male mice is shown). LDL cholesterol is significantly decreased in knockout mice compared with wild-type mice (E) [adjusted P=0.012, male knockout effect estimated: −0.35±0.10 mM (±standard error)]. The wild-type data, collected as local controls (phenotyped in the same week), are shown in the figure as the boxplot, whereas the green band shows the natural variation in the parameter as assessed by the 95% confidence intervals in control mice of that genetic background. Blood fat parameters between WT and knockout mice were compared using a mixed model statistical model. Histopathology revealed absent or minimal hepatic lipidosis in Tbc1d10a−/− mice (F, bottom panel). Marked hepatic lipidosis is evident in wild-type mice (F, top panel).
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f3-0070515: Histopathology complements clinical phenotypes from the primary screen.Lrig−/− mice were annotated to have abnormal skin (data from male mice is shown) (Fisher’s exact test adjusted P-value=2.09e–10). The number and proportion of affected mice in knockout and wild-type baseline controls is presented (A). Abnormal skin with scaly foci (arrows) was observed in Lrig−/− mice (B). Histopathology revealed epidermal hyperplasia (arrow) and hyperkeratosis (arrowhead) (C). Mcph1−/− mice were annotated as infertile. Ovarian hypoplasia with absence of folliculogenesis was observed in Mcph1−/− females (D, top right panel); ovary from a wild-type mouse with growing follicles (arrows) is shown (D, top left panel). Seminiferous tubule vacuolation with lack of germ cells (arrows) was observed in Mcph1−/− males (D, bottom right panel); normal testis with abundant developing germ cells (arrows) is shown from a wild-type mouse (D, bottom left panel). Tbc1d10a−/− mice were annotated to have decreased circulating low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol (LDL data from male mice is shown). LDL cholesterol is significantly decreased in knockout mice compared with wild-type mice (E) [adjusted P=0.012, male knockout effect estimated: −0.35±0.10 mM (±standard error)]. The wild-type data, collected as local controls (phenotyped in the same week), are shown in the figure as the boxplot, whereas the green band shows the natural variation in the parameter as assessed by the 95% confidence intervals in control mice of that genetic background. Blood fat parameters between WT and knockout mice were compared using a mixed model statistical model. Histopathology revealed absent or minimal hepatic lipidosis in Tbc1d10a−/− mice (F, bottom panel). Marked hepatic lipidosis is evident in wild-type mice (F, top panel).

Mentions: In 19 of 30 lines (63.3%) with MGP clinical phenotypes, histopathology revealed lesions that were associated with at least one documented clinical phenotype (Table 1). In 17 of these lines, histopathology revealed an unequivocal pathology phenotype that potentially explained at least one documented clinical phenotype, and added morphologically and biologically relevant information to the phenotype annotation. For example, histopathology in Lrig1tm1a mice revealed epidermal and follicular hyperplasia with hyperkeratosis consistent with abnormal skin (scaly skin) observed in clinical phenotyping (Fig. 3A–C). In some lines, multiple pathology phenotypes were observed, consistent with multiple clinical phenotype hits. A case in point is the Mcph1tm1a line, which showed a range of clinical phenotypes, including male and female infertility, decreased auditory brainstem response, absent pinna reflex, abnormal eye morphology including corneal vascularization, micronuclei, decreased bone mineral content and decreased bone trabeculae (Chen et al., 2013). Histopathology of this line revealed gonadal hypoplasia and absence of gametogenesis in both females and males (consistent with infertility) (Fig. 3D), corneal thickening and hyaloid artery remnant (consistent with abnormal eye morphology), trabecular osteopenia in long bones (consistent with decreased bone trabeculae), and small brain/ microencephaly (consistent with microcephaly) (data not shown). The clinical and pathology phenotypes in this line were broadly consistent with microcephaly, a primary autosomal-recessive human condition associated with mutations of MCPH1 [Online Mendelian Inheritance in Man (OMIM) 251200 (www.ncbi.nlm.nih.gov/omim)] (supplementary material Table S5).


Histopathology reveals correlative and unique phenotypes in a high-throughput mouse phenotyping screen.

Adissu HA, Estabel J, Sunter D, Tuck E, Hooks Y, Carragher DM, Clarke K, Karp NA, Sanger Mouse Genetics ProjectNewbigging S, Jones N, Morikawa L, White JK, McKerlie C - Dis Model Mech (2014)

Histopathology complements clinical phenotypes from the primary screen.Lrig−/− mice were annotated to have abnormal skin (data from male mice is shown) (Fisher’s exact test adjusted P-value=2.09e–10). The number and proportion of affected mice in knockout and wild-type baseline controls is presented (A). Abnormal skin with scaly foci (arrows) was observed in Lrig−/− mice (B). Histopathology revealed epidermal hyperplasia (arrow) and hyperkeratosis (arrowhead) (C). Mcph1−/− mice were annotated as infertile. Ovarian hypoplasia with absence of folliculogenesis was observed in Mcph1−/− females (D, top right panel); ovary from a wild-type mouse with growing follicles (arrows) is shown (D, top left panel). Seminiferous tubule vacuolation with lack of germ cells (arrows) was observed in Mcph1−/− males (D, bottom right panel); normal testis with abundant developing germ cells (arrows) is shown from a wild-type mouse (D, bottom left panel). Tbc1d10a−/− mice were annotated to have decreased circulating low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol (LDL data from male mice is shown). LDL cholesterol is significantly decreased in knockout mice compared with wild-type mice (E) [adjusted P=0.012, male knockout effect estimated: −0.35±0.10 mM (±standard error)]. The wild-type data, collected as local controls (phenotyped in the same week), are shown in the figure as the boxplot, whereas the green band shows the natural variation in the parameter as assessed by the 95% confidence intervals in control mice of that genetic background. Blood fat parameters between WT and knockout mice were compared using a mixed model statistical model. Histopathology revealed absent or minimal hepatic lipidosis in Tbc1d10a−/− mice (F, bottom panel). Marked hepatic lipidosis is evident in wild-type mice (F, top panel).
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Related In: Results  -  Collection

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f3-0070515: Histopathology complements clinical phenotypes from the primary screen.Lrig−/− mice were annotated to have abnormal skin (data from male mice is shown) (Fisher’s exact test adjusted P-value=2.09e–10). The number and proportion of affected mice in knockout and wild-type baseline controls is presented (A). Abnormal skin with scaly foci (arrows) was observed in Lrig−/− mice (B). Histopathology revealed epidermal hyperplasia (arrow) and hyperkeratosis (arrowhead) (C). Mcph1−/− mice were annotated as infertile. Ovarian hypoplasia with absence of folliculogenesis was observed in Mcph1−/− females (D, top right panel); ovary from a wild-type mouse with growing follicles (arrows) is shown (D, top left panel). Seminiferous tubule vacuolation with lack of germ cells (arrows) was observed in Mcph1−/− males (D, bottom right panel); normal testis with abundant developing germ cells (arrows) is shown from a wild-type mouse (D, bottom left panel). Tbc1d10a−/− mice were annotated to have decreased circulating low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol (LDL data from male mice is shown). LDL cholesterol is significantly decreased in knockout mice compared with wild-type mice (E) [adjusted P=0.012, male knockout effect estimated: −0.35±0.10 mM (±standard error)]. The wild-type data, collected as local controls (phenotyped in the same week), are shown in the figure as the boxplot, whereas the green band shows the natural variation in the parameter as assessed by the 95% confidence intervals in control mice of that genetic background. Blood fat parameters between WT and knockout mice were compared using a mixed model statistical model. Histopathology revealed absent or minimal hepatic lipidosis in Tbc1d10a−/− mice (F, bottom panel). Marked hepatic lipidosis is evident in wild-type mice (F, top panel).
Mentions: In 19 of 30 lines (63.3%) with MGP clinical phenotypes, histopathology revealed lesions that were associated with at least one documented clinical phenotype (Table 1). In 17 of these lines, histopathology revealed an unequivocal pathology phenotype that potentially explained at least one documented clinical phenotype, and added morphologically and biologically relevant information to the phenotype annotation. For example, histopathology in Lrig1tm1a mice revealed epidermal and follicular hyperplasia with hyperkeratosis consistent with abnormal skin (scaly skin) observed in clinical phenotyping (Fig. 3A–C). In some lines, multiple pathology phenotypes were observed, consistent with multiple clinical phenotype hits. A case in point is the Mcph1tm1a line, which showed a range of clinical phenotypes, including male and female infertility, decreased auditory brainstem response, absent pinna reflex, abnormal eye morphology including corneal vascularization, micronuclei, decreased bone mineral content and decreased bone trabeculae (Chen et al., 2013). Histopathology of this line revealed gonadal hypoplasia and absence of gametogenesis in both females and males (consistent with infertility) (Fig. 3D), corneal thickening and hyaloid artery remnant (consistent with abnormal eye morphology), trabecular osteopenia in long bones (consistent with decreased bone trabeculae), and small brain/ microencephaly (consistent with microcephaly) (data not shown). The clinical and pathology phenotypes in this line were broadly consistent with microcephaly, a primary autosomal-recessive human condition associated with mutations of MCPH1 [Online Mendelian Inheritance in Man (OMIM) 251200 (www.ncbi.nlm.nih.gov/omim)] (supplementary material Table S5).

Bottom Line: In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen.Three of these seven lines had no clinical phenotype detected by the standard primary screen.Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls.

View Article: PubMed Central - PubMed

Affiliation: Centre for Modeling Human Disease, Toronto Centre for Phenogenomics, 25 Orde Street, Toronto, ON M5T 3H7, Canada.

ABSTRACT
The Mouse Genetics Project (MGP) at the Wellcome Trust Sanger Institute aims to generate and phenotype over 800 genetically modified mouse lines over the next 5 years to gain a better understanding of mammalian gene function and provide an invaluable resource to the scientific community for follow-up studies. Phenotyping includes the generation of a standardized biobank of paraffin-embedded tissues for each mouse line, but histopathology is not routinely performed. In collaboration with the Pathology Core of the Centre for Modeling Human Disease (CMHD) we report the utility of histopathology in a high-throughput primary phenotyping screen. Histopathology was assessed in an unbiased selection of 50 mouse lines with (n=30) or without (n=20) clinical phenotypes detected by the standard MGP primary phenotyping screen. Our findings revealed that histopathology added correlating morphological data in 19 of 30 lines (63.3%) in which the primary screen detected a phenotype. In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen. Three of these seven lines had no clinical phenotype detected by the standard primary screen. Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls. These findings demonstrate the complementary and unique contribution of histopathology to high-throughput primary phenotyping of mutant mice.

Show MeSH
Related in: MedlinePlus