Background and Observations

• This study investigates how left-right (LR) asymmetry is established during chick embryogenesis long before any visible physical differences appear.
• The research focuses on the expression patterns of key genes that help determine the proper positioning of internal organs such as the heart.
• The main genes examined are an activin receptor (with two forms), Sonic hedgehog (Shh), and a nodal-related gene (cNR-1).

Key Genes Involved

• Activin Receptors:
  • cAct-RIIb is expressed symmetrically in the primitive streak and Hensen’s node.
  • cAct-Rlla is expressed asymmetrically, with stronger expression on the right side of Hensen’s node. It is inducible by activin protein.
• Sonic hedgehog (Shh):
  • Initially expressed symmetrically in Hensen’s node at early stages.
  • Later, its expression becomes restricted to the left side, serving as a key signal for left-right patterning.
• cNR-1 (Nodal-related):
  • Begins with a subtle left-sided expression near Hensen’s node during early stages.
  • Later, a larger expression domain appears in the lateral plate mesoderm, which contributes to the formation of heart tissue.

Observations of Asymmetric Gene Expression

• Researchers used in situ hybridization to visualize gene expression in chick embryos from stage 4 to stage 7.
• While many genes show symmetric expression, cAct-Rlla, Shh, and cNR-1 exhibit clear left-right differences in Hensen’s node and surrounding tissues.
• Cryosectioning confirmed that the asymmetry exists in specific tissue layers, such as the ectoderm versus the mesoderm.

Experimental Manipulations and Their Effects

• Activin Bead Implants:
  • Implanting an activin-soaked bead on the left side of Hensen’s node induced abnormal (ectopic) expression of cAct-Rlla on that side.
  • This treatment also repressed the normal expression of Shh on the left side, altering the usual asymmetry.
• Shh Cell Pellet Implants:
  • When cell pellets expressing Shh were implanted on the right side, an ectopic domain of cNR-1 expression appeared on that side.
  • This result shows that Shh acts upstream in the cascade by inducing cNR-1 expression.
• Effects on Heart Laterality:
  • Exposure of embryos to either activin or Shh on both sides resulted in a randomization of heart orientation (heart situs).
  • Normally, the heart loops to the right, but manipulation of these signals can invert this process, demonstrating their role in determining organ positioning.

Molecular Cascade Model for LR Asymmetry

• The proposed model suggests an early, sequential signaling cascade that establishes LR asymmetry:
• An asymmetrically distributed activin-like signal (stronger on the right) induces cAct-Rlla expression in the right side of Hensen’s node.
• This, in turn, restricts Shh expression to the left side of the node.
• Shh then signals to adjacent cells, inducing cNR-1 expression in the lateral plate mesoderm, which contains cardiac precursor cells.
• In simple terms, these steps work together like following a recipe, where each molecular signal triggers the next step in establishing left-right differences.

Conclusions and Implications

• The study demonstrates that the chick embryo establishes left-right asymmetry through a cascade of gene interactions well before any visible asymmetry appears.
• Manipulating these signals can alter the normal positioning of the heart, highlighting their crucial role in proper organ development.
• This research provides insight into the molecular basis of LR asymmetry, which may help in understanding congenital defects related to organ placement.

背景与观察 (引言)

• 本研究探讨了鸡胚胎在出现明显形态差异之前，如何建立左右（LR）不对称性。
• 研究重点在于几个关键基因的表达模式，这些基因决定了心脏等内脏器官的正确位置。
• 主要研究基因包括激素受体（有两种形式）、Sonic hedgehog (Shh) 以及与Nodal相关的基因 cNR-1。

关键基因

• 激素受体：
  • cAct-RIIb 在原始条和亨森结节中对称表达。
  • cAct-Rlla 在亨森结节右侧表达较强，其表达可被激素诱导。
• Sonic hedgehog (Shh)：
  • 初期在亨森结节中对称表达。
  • 随后其表达局限于左侧，起到左右模式形成的关键信号作用。
• cNR-1 (Nodal相关)：
  • 在早期阶段，cNR-1在结节附近左侧有细微表达。
  • 之后在侧板中出现较大区域的表达，这部分组织包含未来心脏的前体细胞。

不对称基因表达的观察

• 研究人员利用原位杂交技术，在鸡胚胎4期到7期观察基因表达情况。
• 结果显示，尽管许多基因表达呈对称分布，但 cAct-Rlla、Shh 和 cNR-1 在亨森结节及其周围组织中表现出明显的不对称性。
• 冷冻切片进一步确认了这种不对称性存在于特定组织层，如外胚层与中胚层之间。

实验操作及其效果

• 激素珠植入：
  • 在结节左侧植入浸泡激素的珠子后，该区域出现了异常的 cAct-Rlla 表达。
  • 同时，左侧 Shh 的正常表达受到抑制，改变了原有的不对称模式。
• Shh 细胞团植入：
  • 在结节右侧植入表达 Shh 的细胞团，导致右侧出现异常的 cNR-1 表达区域。
  • 这表明 Shh 位于信号级联的上游，能够诱导 cNR-1 的表达。
• 心脏位向变化：
  • 当胚胎同时暴露于激素或 Shh 时，心脏的左右位置（心脏位向）会变得随机化。
  • 通常情况下，心脏向右弯曲，但干预这些信号会导致该过程发生反转，证明了这些分子在器官定位中的关键作用。

左右不对称的分子级联模型

• 模型提出胚胎早期存在一个依次进行的信号级联来建立左右不对称性：
• 首先，在右侧存在较强的激素样信号，通过诱导 cAct-Rlla 在亨森结节右侧表达起作用。
• 这种激素诱导效应限制了 Shh 只能在结节左侧表达。
• Shh 随后作为信号，诱导邻近侧板中 cNR-1 的表达，而侧板中正包含未来形成心脏的细胞。
• 简单来说，这一过程就像按照食谱一步步操作，各分子依次触发下一步，从而建立左右不对称性。

结论与意义

• 研究表明，鸡胚胎在形态上出现不对称之前，就已通过一系列基因相互作用建立了左右不对称性。
• 干预这些信号能够改变心脏的正常定位，证明它们在器官发育中起着关键作用。
• 该研究为揭示左右不对称的分子机制提供了重要线索，对理解与器官定位相关的先天性缺陷具有重要意义。