Cilia-related diseases.

There are at least eight categories of cilia in the human body and malfunctioning of any one or several of them will have different consequences for the patient. A genetic error of the respiratory cilia (9 + 2) is the cause of the airways disease named immotile-cilia syndrome (or PCD), whereas defective ependymal cilia (9 + 2) carries an increased risk of hydrocephalus. When the so-called nodal cilia (9 + 0) of the early embryo are malfunctioning, there is a random determination of asymmetry of the heart and visceral organs ('a 50% risk of situs inversus'). Some genes are responsible for the synthesis, transport, and assembly of the cilia, and mutations in these genes may lead to progressive degeneration of ciliary structures, such as the connecting cilium (9 + 0) of the photoreceptor cells-this is the cause of retinitis pigmentosa. Ciliary malfunctions due to genetic errors tend to be systemic and life-long, whereas acquired diseases are local and may be temporary only.

Sperm structure and spermiogenesis in Coletinia sp. (Nicoletiidae, Zygentoma, Insecta) with a comparative analysis of sperm structure in Zygentoma.

The spermatozoon of Coletinia sp. has a bilayered acrosome, a short nucleus (4 microm) and a relatively short sperm tail with two mitochondrial derivatives. The chromatin is uniformly dense except for several electron-lucid channels or strands which permeate the nucleus and which originate in the spermatid as invaginations of the nuclear envelope. The invaginations occur mostly or exclusively along two meridians of the spermatid that are also characterized by the presence of a longitudinal rod of medium electron density. The two rods (designated as 'mid-spermatid rods') evidently are instrumental in the formation of the electron-lucid channels. The significance of this elaborate system of intranuclear channels is not understood. The sperm tail has a 9 + 9 + 2 axoneme with each of the nine microtubular doublets accompanied by an accessory microtubule; scant intertubular material can also be distinguished. Hence, the tail axoneme resembles that of many pterygote insects. Each of the two mitochondrial derivatives contains a crystalline inclusion that has periodically spaced layers going in different directions on either side of the midline. Two synapomorphic traits appear to be shared by Ateluridae and Nicoletiidae, namely the invaginations of the nuclear membrane along two meridians of the nucleus and the shape of the crystalline inclusions of the mitochondrial derivatives. Four species from the family Lepismatidae were also examined as to their sperm ultrastructure. Three of them, Allacrotelsa kraepelini, Ctenolepisma longicaudata and Ctenolepisma sp., were found to be very similar to the two previously examined lepismatids, Thermobia domestica and Lepisma saccharina. On the other hand, spermatozoa of Tricholepisma aurea were aggregated in small groups rather than pairwise joined as seen in the other lepismatids. Sperm characters are also used to reconstruct a phylogenetic hypothesis which suggests a close relationship between Ateluridae and Nicoletiidae.

New findings on sperm ultrastructure of Xenos vesparum (Rossi) (Strepsiptera, Insecta).

The systematic position of insect order Strepsiptera is still under debate. It was, therefore, thought of interest to examine the ultrastructure of a strepsipteran in a search for synapomorphies shared with Coleoptera, Diptera, or any other insect order. The fine structure of spermatozoa and the spermatid from Xenos vesparum (Rossi) was re-examined using scanning and transmission electron microscopy and a fixation technique that permits the visualization of the macromolecular organization of the organelles. The spermatozoon was shown to possess several traits that are characteristics of insects in general, such as a 9 + 9 + 2 axoneme, two mitochondrial derivatives containing a crystalline material and two 'zipper lines' present along the sperm tail. Seventeen protofilaments occurred along most of the accessory tubules, which reduced to 16 posteriorly. An acrosome is absent. The neck region contains a prominent centriolar adjunct, which gives rise to two accessory bodies which adhere to the mitochondrial derivatives, and to slender strands of the so-called intertubular material found between the accessory tubules. Of interest is the finding that the glycocalyx consists of prominent filamentous strands, similar to those found in siphonapterans, mecopterans and basal dipterans.

Sperm ultrastructure and spermiogenesis in the relic species Tricholepidion gertschi Wygodzinsky (Insecta, Zygentoma).

The silverfish Tricholepidion gertschi is of interest in that it is the most basal representative of Zygentoma. An ultrastructural study of its spermiogenesis was performed to find out whether there are traits which resemble those of other, more advanced insects. This was found to be the case; spermiogenesis can be considered to be of a common insectan type, leading to the formation of elongated sperm cells with acrosome, nucleus, neck region and a tail with axoneme and two mitochondrial derivatives. Total cell length, 50 microm, is short for an insect. There are some specializations, which probably represent autapomorphies. The acrosome has a posterior canal or cleft that makes a U-turn. The centriole adjunct forms a prominent post-nuclear ring surrounding the centriole and have a posterior extension, and further originates nine intertubular fibers with a longitudinal periodicity and two accessory bodies. The mitochondrial derivatives have five rows of regularly spaced cristae within a crystalline matrix. The axoneme has accessory tubules consisting of 16 protofilaments, formed at the B-tubules of the doublets and placed at some distance from them in the posterior part of the sperm tail.

Binucleate and biflagellate spermatozoa in Tricholepidion gertschi Wygodzynsky (Insecta, Zygentoma).

The phenomenon of sperm pairing is known from some species of the apterygotan insect order Zygentoma, and has been described as the close apposition of two sperm cells. When released from the testes, they are single cells; pairing taking place in the deferent ducts. In a study of the relic species Tricholepidion gertschi, Zygentoma, sperm pairing was found to be due to a true fusion of two partners along their entire sperm head regions. The spermatozoon thus formed has two acrosomes, two nuclei and two separate sperm tails. The biflagellate spermatozoon swims with coordinated movements of its two flagella only when the two flagella lie close together but is totally uncoordinated when separate. The spermatozoon is about 50 microm long, thus much shorter than those of related apterygotan species. The mechanism of sperm cell fusion is unclear, although it appears that a 55-nm wide layer of electron dense substance, here termed the peripheral lamina, may play a role in delimiting the extent of sperm fusion.

Ciliary structure in health and disease.

The architecture of the cilium seemingly is very simple but the appearance is deceptive; its 'engine' actually is quite complex. There is a circle of nine microtubular doublets around two central singlet microtubules, with dynein arms and spokes extending from the doublets, and nexin links joining them. The inner microtubules are partly surrounded by a central sheath. Many uncertainties remain, for instance the role of the spokes, the nexin links and of the various categories of inner dynein arms. The complexity as seen today is largely the result of new methodologies being introduced, such as the deep-etch technique or image processing using various computer programs. The inborn disease named immotile-cilia syndrome is characterized by the cilia being defective. It is a highly heterogeneous disease in that more than a dozen subgroups characterized by different ciliary defects have been recognized.

Spermatozoa of the 'primitive type' in Scutigerella (Myriapoda, Symphyla).

The myriapod class Symphyla is of interest in that insects generally are assumed to be derived from symphylan-like ancestors. Male Symphyla form spermatophores that are picked up by the female. Both euspematozoa and paraspermatozoa are formed. In spite of the mode of fertilization their euspermatozoa were found to be of a kind that is typical of aquatic animals, so called 'primitive spermatozoa;' these are characterized by a short sperm head with a bilayered acrosome, a midpiece containing a few unmodified mitochondria, and a 9 + 2 flagellum. Scutigerella are unique among terrestrial arthropods in having 'primitive spermatozoa'; and together with horseshoe crabs they are only the second case in Arthropoda. Two further sperm plesiomorphies not found in other myriapods or insects are (1) the presence of a cytoplasmic canal housing the proximal flagellum and (2) the existence of microtubular triplets in centrioles, one of which acts as a basal body. Symphyla and Diplopoda both have a striated structure in the center of the subacrosomal material. The paraspermatozoa lack acrosome and nucleus but have a prominent crystal, a single mitochondrion, and two membrane systems. The structure of Scutigerella euspermatozoa is consistent with Symphyla being close to the stem group of Myriapoda plus Insecta.

Primary ciliary dyskinesia: a genome-wide linkage analysis reveals extensive locus heterogeneity.

Primary ciliary dyskinesia (PCD), or immotile cilia syndrome (ICS), is an autosomal recessive disorder affecting ciliary movement with an incidence of 1 in 20000-30000. Dysmotility to complete immotility of cilia results in a multisystem disease of variable severity with recurrent respiratory tract infections leading to bronchiectasis and male subfertility. Ultrastructural defects are present in ciliated mucosa and spermatozoa. Situs inversus (SI) is found in about half of the patients (Kartagener syndrome). We have collected samples from 61 European and North American families with PCD. A genome-wide linkage search was performed in 31 multiplex families (169 individuals including 70 affecteds) using 188 evenly spaced (19cM average interval) polymorphic markers. Both parametric (recessive model) and non-parametric (identity by descent allele sharing) linkage analyses were used. No major locus for the majority of the families was identified, although the sample was powerful enough to detect linkage if 40% of the families were linked to one locus. These results strongly suggest extensive locus heterogeneity. Potential genomic regions harbouring PCD loci were localised on chromosomes 3p, 4q, 5p, 7p, 8q, 10p, 11q, 13q, 15q, 16p, 17q and 19q. Linkage analysis using PCD families with a dynein arm deficiency provided 'suggestive' evidence for linkage to chromosomal regions 8q, 16pter, while analyses using only PCD families with situs inversus resulted in 'suggestive' scores for chromosomes 8q, and 19q.

Movement of turritella spermatozoa: direction of propagation and chirality of flagellar bends.

The marine snail, Turritella communis, produces two types of spermatozoa, named apyrene and eupyrene. Eupyrene spermatozoa are usually paired, but unpaired ones are involved in fertilization. Movements of these spermatozoa were analyzed using a video camera with a high-speed shutter. The eupyrene spermatozoa usually swim with the head foremost but are able to swim flagellum foremost. A reversal of the direction of their swimming was found to be the result of a change in the direction of flagellar bend propagation, which changed with calcium concentration. Reversal of the direction of bend propagation was accompanied by a reversal of direction of the rotational movement of the spermatozoa around their long axis, suggesting that the bending waves keep the sense of their three-dimensional form. The swimming speed of apyrene spermatozoa in natural seawater was about one-eighth of that of the eupyrene ones and remained almost constant in highly viscous medium. The swimming speed of conjugated eupyrene spermatozoa was the same as that of unpaired spermatozoa over a wide viscosity range (<3,000 cP). No advantage of swimming by two spermatozoa could be detected in Turritella spermatozoa.

Asymmetry of cilia and of mice and men.

Evidence is given for the opinion that cilia in the early embryo, by their work, determine the laterality of the body; without ciliary work body laterality would be randomized. More exactly, monocilia in the primitive node are responsible for this determination. They have been described as being of the 9+0 type, but with dynein arms and with a gyrating movement. The orientation of the monocilia on the epithelium is of no importance but the direction of their gyration is, as may also be the shape of the node. The chirality of the cilia is thus reflected directly in the asymmetry of the body. The dynein arms go clockwise as seen from the base to tip and the ciliary rotation is in the same direction. The resulting waterflow is towards the left and so is the movement of the forming heart. In most subgroups of the immotile-cilia syndrome this mechanism does not work and equally many individuals will be born with situs inversus as with situs solitus. An exception is the immotile-cilia subgroup, named 'microtubule transposition', which is characterized by all cilia having a 9+0 structure throughout most of their length.

Inheritance of randomness.

The mouse mutant iv is characterized by "a random determination of a developmental process' in that 50 rather than 100% of the homozygotes have situs inversus. The same explanation is given to the inheritance of situs inversus in the human immotile-cilia syndrome. There are probably two alleles of the responsible gene, one for control of the proper asymmetry and one without control and hence resulting in equal numbers of situs solitus and situs inversus in the homozygote. Left-handedness may be similarly inherited; furthermore, because of its high prevalence (around 10-12%) it has been assumed that there is an advantage of carrying both alleles ("balanced polymorphism'). With these two assumptions, a prevalence of left-handedness of 12.5% is expected, a 50% chance of left-handedness in matings between two left-handed persons, and equal numbers of discordant and left-handed concordant monozygotic twins. These values are close to those actually found. As the values for (female) homosexuality are similar to those of left-handedness a similar inheritance is proposed.

The sperm structure of the gall-midges Anaretella and Lestremia (Insecta, Diptera, Cecidomyiidae).

The sperm structure of some dipteran flies belonging to the Lestremiini tribe have been examined. Anaretella cincta was shown to have an axoneme made of 20-21 microtubular doublets, disposed in a circle in a cross section and surrounding a mitochondrion. Other crystal-containing mitochondria flank the axoneme; a second species (Anaretella sp.) was provided with 21-22 axonemal doublets. Lestremia is characterized by a flattened axoneme, consisting of about 150 doublets arranged in 2 antiparallel rows and surrounding a few mitochondria. These mitochondria, in Lestremia sp., have a crystalline core that is missing in Lestremia cinerea. The structure of microtubular doublets is quite similar in the 2 related genera and a derivation of the flattened axoneme found in Lestremia from that circular of Anaretella is suggested. Sperm structure suggests that Lestremia cinerea is not a uniform species.

Situs inversus and ciliary abnormalities. What is the connection?

The finding of men with living but immotile sperm tails has initiated a search for the cause of the disorder. The sperm tails were found to lack dynein arms or to have some other ultrastructurally visible defect and the cilia were found to have the same defects. The disorder was hence named the immotile-cilia syndrome. Two more groups with the same clinical symptoms were later found, characterized by ciliary dysmotility or ciliary aplasia. In each group there are several subgroups. Many of the affected persons have situs inversus totalis; in some subgroups the incidence of situs inversus is probably 50%; there is, thus, a random determination of visceral asymmetry. Five hypotheses have been forwarded that attempt to explain the connection between ciliary defects and loss of laterality control. Support for, or evidence against, these five hypotheses have been sought in some animal models of the syndrome. Whereas immotile-cilia syndrome in dogs and pigs is very similar to the human one, an animal model in the rat differs from the human syndrome in that mainly the males are affected. Two animal models in the mouse differ in that one has ciliary defects but no increased incidence of situs inversus and the other has a random determination of visceral laterality and no ciliary defects. The connection between ciliary defects and random determination of laterality remains enigmatic.

Flagellar structure in normal human spermatozoa and in spermatozoa that lack dynein arms.

Human sperm flagella were analyzed by electron microscopy and computer averaging in order to characterize normal flagella and to detect differences between normal and mutated spermatozoa. The A-tubules of normal spermatozoa were seen to have 13 protofilaments and a lumen containing a 'pentagon' and a 'sickle'. The incomplete B-tubule was seen to have 10 protofilaments with an angular separation such that a complete circle would have 16 protofilaments. A thin '11th filament' is located at the inner border between A- and B-tubules and, in the centriole, also between B- and C-tubules. The tail end piece has 18 microtubules of a conventional appearance. We conclude that the 9 axonemal doublets split distally into 2 microtubules and that normal microtubules with 13 protofilaments can grow from the incomplete B-tubules. The cell membrane in the end piece has a glycocalyx with regular periodicity. Spermatozoa from a man suffering from the immotile-cilia syndrome was also analyzed. His sperm flagella were seen to be abnormal in that the dynein arms are lacking, and, that the sickle is incomplete. In other respects his immotile spermatozoa were normal; spokes and central projections have the same appearance as in normal spermatozoa.

Effects of spermidine on function and ultrastructure of alveolar macrophages.

It is known that spermidine and other naturally occurring polyamines accumulate in rat lung slices by an energy-dependent uptake process and that alveolar macrophages (AM) have a greater rate of uptake than has the total lung cell population. In the present study rat AMs were incubated with spermidine, which resulted in a marked and significant (P < 0.002) decrease in phagocytosis of heat-killed yeast cells at concentrations 0.2 and 0.5 mM and a tendency to decrease at 0.05 mM. The number of microtubules surrounding the centrioles was measured using electron microscopy and appeared to be decreased at concentrations 0.2 and 0.5 mM. There was no affect on phagolysosomal pH. The results suggest that spermidine might affect the defense against inhaled pollutants and microbes, especially when spermidine levels are increased, as they are under conditions with high mitotic activity, e.g., in tumors.

Ultrastructure of human nasal epithelium during an episode of coronavirus infection.

The nasal epithelium from a young girl was examined by electron microscopy and found to be infected by coronavirus. Virions are seen within and outside the ciliated cells, but not outside or within the goblet cells or other cells of the nasal mucosa. Some virions are located near the microvilli, others in pockets in the apical cell membrane. The cytoplasm contains many small vesicles with a single virion, large apical vesicles containing hundreds of virions, and lysosome-like cytosomes with a moderate number of virions. Some virus-like particles devoid of an electron-dense interior are seen both in the cytosomes and extracellularly. Virus budding was observed in the Golgi apparatus but nowhere else in the cell. The ciliated cells seem not to be destroyed by the viruses, although in many cases the cilia are withdrawn into the cell body. The loss of cilia is likely to cause rhinorrhoea.

Movement of Myzostomum spermatozoa: calcium ion regulation of swimming direction.

Spermatozoa of the small myzostomid worm Myzostomum cirriferum usually swim with the flagellum foremost but occasionally stop and then swim with the head foremost. The spermatozoa have axoneme of the 9 + 0 type; thus each lacks the central pair microtubules. The flagellum emerges in the anterior end of the cell body and attaches to it with junctions. To understand the mechanism regulating the swimming direction of the spermatozoa, we recorded the sperm and their flagellar movements using a video camera with a high-speed shutter. The effects of calcium and viscosity on these movements were also examined. The cell body with the flagellum attached to it formed a curved plate during beating, while the free portion of the flagellum beats with small helical bends. Motive force to propel a spermatozoon was mainly due to the bends in the cell body. The spermatozoa reversed the direction of their swimming as a result of a change in the direction of bend propagation. The direction of bend propagation was regulated by calcium; the bends in the cell body propagated from the end of the head toward the free portion of the flagellum at low concentrations of Ca2+, whereas the direction of bend propagation was reversed at high concentrations of this ion. High viscosity of the medium stimulated a change in the direction of bend propagation.