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Figure 4. Depletion of lhx1 results in a loss of the pronephric kidney.Embryos were injected (1xV2) with 300 pg of ctrl-AS or lhx1-AS at the 8-cell stage. (A, D, G, J) Uninjected embryos. (B, E, H, K) Embryos injected with ctrl-AS. (C, F, I, L) Embryos injected with lhx1-AS. (A–C) In situ hybridization of embryos at stage 20 for the pronephric marker pax8. (C) Reduced or absent pax8 expression was observed in 47% and 53% of the embryos, respectively (n = 34). (D–F) 3G8 whole-mount immunostaining was carried out at stage 32. (E) Reduced 3G8 staining was observed in 5% of the embryos (n = 41). (F) Reduced 3G8 staining was observed in 76% of the embryos (n = 34). Insets show enlargement of 3G8 staining of the proximal tubule. (G–I) 4A6 whole-mount immunostaining was carried out at stage 37/38. (H) Reduced 4A6 staining was observed in 9% of the embryos (n = 23). (I) Reduced 4A6 staining was observed in 91% of the embryos (n = 34). (J–L) In situ hybridization of embryos at stage 39 for β1-NaK-ATPase. (L) Reduced β1-NaK-ATPase expression was observed in 72% of the embryos (n = 29). (M) Bar graph with the percentage embryos injected with the different doses of lhx1-AS that showed presence, reduction or absence of pax8 expression at stage 20. Lhx1-AS 100 pg: reduced pax8 expression was observed in 89%, absence in 4% and presence in 7% of the embryos (n = 28). Lhx1-AS 200 pg: reduced pax8 expression was observed in 93% and presence in 7% of the embryos (n = 27). Lhx1-AS 300 pg: reduced of pax8 expression was observed in 47% and absent in 53% of the embryos (n = 34). Lhx1-AS 400 pg: reduced pax8 expression was observed in 50% and absent in 50% of the embryos (n = 42). Lhx1-AS 300 pg+zflhx1 mRNA 25 pg: presence of pax8 expression was observed in 51% of the embryos (n = 41). |