Groundbreaking 3D imaging unveils secrets of cilia

Primary cilia are sensory organelles present on the cell surface; however, their physical structure has not been defined due to technical reasons. A new PNAS study examined primary cilia in human islet cells using a multi-scale surface scanning electron microscope approach to obtain a three-dimensional (3D) image of their architecture.

Study: Scanning electron microscopy of human islet cilia. Image Credit: Jose Luis Calvo / Shutterstock.com Study: Scanning electron microscopy of human islet cilia. Image Credit: Jose Luis Calvo / Shutterstock.com

Introduction

Human islet primary cilia are involved in the regulation of blood glucose levels in the body. These organelles arise from the centriole of the cell, which forms the ciliary basal body, and extend into the extracellular space. When the function of cilia is impaired or their number is reduced, metabolic disease arises due to altered pancreatic development.

Pancreatic islet primary cilia were first identified on beta cells, with each cell having one cilium. These cilia consist of multiple subdomains, with each cilium having a basal body and axoneme. The axoneme has a characteristic nine-doublet outer ring of microtubules, with no central microtubule at the base; however, this structure may change after cilia emerge from the cell surface.

Scanning electron microscopy (SEM) is used to study the shape of surface structures; however, this approach sacrifices submembrane architecture during sample preparation. Nevertheless, advanced microscopy and analytics technology may help examine the structure of primary cilia along the entire length.

In the current study, scientists couple SEM with membrane extraction to delineate the surface morphology of primary cilia, along with the structure of axonemes beneath the ciliary membrane. Some cilia that had their membranes removed were subsequently examined to define the number and size of microtubules, other structural components repeating in such cilia, and chirality.

Human islet cells in the pancreas sport long antenna-like organelles called cilia that sense glucose. A new imaging study provides a detailed look at these vital structures. In PNAS: https://t.co/4Iwzm6FjYa pic.twitter.com/AuYNc0NLWB

— PNASNews (@PNASNews) May 29, 2023

What is the ciliary structure?

The researchers viewed the primary cilia as prominent surface structures between three and six micrometers (μm) long. Intact cilia were imaged to determine the structure of the axoneme throughout their length. Correspondingly, demembranated cilia were imaged to examine the whole axonemal structure.

Most cilia were directed away from the cell and estimated to be about six μm long. The average basal diameter was 240 nanometers (nm) wide at the base and 50 nm at the tip. These measurements corroborate those of earlier studies.

Each cilium had a volume of about 0.15 μm3, which is 5,000 times less than the corresponding cell. Furthermore, each cilia had a surface area of 3.05 μm2, which is about 200 times less than the cell's plasma membrane. Each cilium arises from a ciliary pocket of the plasma membrane, which performs multiple anchoring and ciliogenesis functions.

The ciliary pocket varied between the different cell types in the islet, thus indicating distinct membrane organization at the base of the cilia in each cell type. Some cilia exhibited a pit-shaped ciliary pocket, whereas others had very deep pockets or no visible pocket.

Moving upwards along the ciliary length, a ciliary necklace, which comprised five to six rows of particles around the base perpendicular to the microtubule direction, was identified. The necklace is about 230 nm in diameter and 140 nm in height. These particles may be key to the transition from triplets to doublets, thereby allowing membrane diffusion to occur.

The base of each cilium appears tethered by fine threads to the microvilli and cytoskeleton surface. These threads resemble actin fibers and are different from the Y-links found in the transition zone immediately following the base.

These asymmetric strands attach to target structures and potentially to the cell surface, thus providing a foundation for the ciliary base. Microtubule doublets change to singlets at about one-third to half the way along the course of the cilium.

As the transition from doublets to singlets occurs, it is accompanied by a helical configuration that is typically left-handed. The rotation arc is about 1,000 nm long, with a pitch between 500-1,000 nm.

This coiling may increase the stability of the microtubule bundle and facilitate ciliary mobility, thereby absorbing shocks and modulating stretch and length functions. However, further studies are needed to confirm these findings, which may help lengthen the axoneme without having to synthesize more microtubules.

The bundle of microtubules appeared to be held together by a submembrane ciliary ring in some beta-cell cilia. The ciliary tip exhibited a distinct organization, with fewer microtubules present towards the distal end. The cilium ends in a dense cap.

The ciliary tip ranges in shape from pointed to bulbous, which may indicate different roles for cilia in different islet cells. In rare cases, a single cell had more than one cilium; however, the purpose of compound cilia remains unknown. These organelles may arise due to planned differential gene expression, maintaining the desired ciliary density, or in response to hormonal factors.

What are the implications?

The study marks the first time human primary cilia have been described in extensive detail. In addition, the ability to examine 60 cilia over their full length provides a reference for future studies in this field.

Here, the ciliary ring was described for the first time in mammals, which may contribute to stability and regulate ciliary assembly.

Further research is needed to understand how ciliary morphology varies between the various islet cell types. The role of actin in ciliary movement, including hypotheses such as the regulation of ciliogenesis and endocytosis, mechanical stabilization of the axoneme, or division of the cilium into functional segments, should also be further investigated.

A 3D architectural study of human primary cilia, the first in pancreatic islets and in any human tissue. Results provide a morphometric basis for understanding ciliary function including signaling capacity, axonemal motility, and cell-cell communication.”

Journal reference:
  • Polino, A. J., Sviben, S., Melena, I., et al. (2023). Scanning electron microscopy of human islet cilia. PNAS. doi:10.1073/pnas.2302624120.
Dr. Liji Thomas

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Dr. Liji Thomas

Dr. Liji Thomas is an OB-GYN, who graduated from the Government Medical College, University of Calicut, Kerala, in 2001. Liji practiced as a full-time consultant in obstetrics/gynecology in a private hospital for a few years following her graduation. She has counseled hundreds of patients facing issues from pregnancy-related problems and infertility, and has been in charge of over 2,000 deliveries, striving always to achieve a normal delivery rather than operative.

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