The promise and perils of stem cell therapeutics

Stem cells are the seeds of tissue repair and regeneration and a promising source for novel therapies. However, apart from hematopoietic stem cell (HSC) transplantation, essentially all other stem cell treatments remain experimental. High hopes have inspired numerous clinical trials, but it has been difficult to obtain unequivocal evidence for robust clinical benefit. In recent years, unproven therapies have been widely practiced outside the standard clinical trial network, threatening the cause of legitimate clinical investigation. Numerous challenges and technical barriers must be overcome before novel stem cell therapies can achieve meaningful clinical impact.     

The promise and perils of exhaled breath condensates

The exhaled breath condensate (EBC) approach provides a convenient and noninvasive approach for sampling the pulmonary epithelial lining fluid (ELF). Increased EBC concentrations of more than a dozen inflammatory markers and hydrogen ions have been reported in lung diseases associated with inflammation. However, the usefulness of EBC is compromised by uncertainties concerning the sources of the EBC droplets and by the extreme and variable dilution of ELF droplets with condensed water vapor ( approximately 20,000-fold). Reported increases in EBC concentrations may reflect proportionate increases in the total volume rather than the concentration of ELF droplets in the collected samples. Conclusions regarding ELF concentrations can only be made if this dilution is estimated with a dilutional indicator (e.g., conductivity of lyophilized EBC). In normal EBC samples, pH is effectively set by oral contamination with NH(3), and EBC pH cannot provide reliable information regarding ELF pH in normal subjects. Acidification of EBC observed in asthma and other conditions may reflect acidification of ELF, decreases in NH(3) added to the EBC, and/or the presence of gastric droplets in the EBC.     

Drinking in Antarctic fishes

Drinking rates have never been measured in Antarctic fish. Drinking rates were measured in four species of notothenioid fish, including a hemoglobinless icefish, found in the near-freezing waters of the Ross Sea of the Southern Ocean. All of the fish, with the exception of the icefish, had low drinking rates and high serum osmolalities relative to temperate seawater fish. The icefish had significantly higher drinking rates and serum osmolalities relative to the Antarctic fishes containing hemoglobin, including Trematomus bernacchii. Warm acclimation of T. bernacchii, from −1.5°C to +4°C for 4 weeks, significantly increased their drinking rates 4.6-fold, significantly decreased their serum and intestinal osmolality by 11% and 12%, respectively, relative to cold-acclimated fish. These results indicate that increased drinking rates in Antarctic fish at elevated temperatures are involved in maintaining a lower serum osmolality.     

The hemoglobin system of Antarctic and non-Antarctic notothenioid fishes

Studies of the hemoglobin system of fish of the suborder Notothenioidei have been extended to non-Antarctic species of Pseudaphritis urvillii and Notothenia angustata. The two species belong to families that were the first to diverge within the suborder. The degree of amino acid sequence identity with Antarctic notothenioids and other non-Antarctic fish species is analyzed with respect to phyletic and ecological diverence.     

The nature of the diversity of Antarctic fishes

The species diversity of the Antarctic fish fauna changed notably during the 40 million years from the Eocene to the present. A taxonomically restricted and endemic modern fauna succeeded a taxonomically diverse and cosmopolitan Eocene fauna. Although the Southern Ocean is 10% of the worlds ocean, its current fish fauna consists of only 322 species, small considering the global diversity of 25,000–28,000 species. The fauna is reasonably well-known from a taxonomic perspective. This intermediate designation between poorly known and well-known indicates that new species are regularly being described. A conservative estimate of the number of undescribed species is 30–60; many of these may be liparids. On the Antarctic continental shelf and upper slope the fauna includes 222 species from 19 families of benthic fishes. The most speciose taxa are notothenioids, liparids and zoarcids, accounting for 88% of species diversity. Endemism for Antarctic species is also, coincidentally, 88%, at least threefold higher than in faunas from other isolated marine localities. Eight notothenioid families, including five that are primarily Antarctic, encompass a total of 44 genera and 129 species, 101 Antarctic and 28 non-Antarctic. The 101 Antarctic species make up 45% of the benthic species diversity in the Antarctic region. However, at the highest latitudes, notothenioids contribute 77% of the species diversity, 92% of the abundance and 91% of the biomass. Although species diversity is low compared to other shelf habitats, the nature of the adaptive radiation in organismal diversity among notothenioids is noteworthy in the marine realm. In some notothenioid clades phyletic diversification was accompanied by considerable morphological and ecological diversification. The exemplar is the benthic family Nototheniidae that underwent a habitat or depth related diversification centred on the alteration of buoyancy. They occupy an array of pelagic and benthopelagic habitats at various depths on the shelf and upper slope. Diversification in buoyancy is the hallmark of the nototheniid radiation and, in the absence of swim bladders, was accomplished by a combination of reduced skeletal mineralisation and lipid deposition. Although neutral buoyancy is found in only five species of nototheniids some, like Pleuragramma antarcticum, are abundant and ecologically important. Much work remains to be done in order to frame and to use phylogenetically based statistical methods to test hypotheses relating to the key features of the notothenioid radiation. To reach this analytical phase more completely resolved cladograms that include phyletically basal and non-Antarctic species are essential.     

Microtubule-associated proteins from Antarctic fishes

Microtubules and presumptive microtubule-associated proteins (MAPs) were isolated from the brain tissues of four Antarctic fishes (Notothenia gibberifrons, N. coriiceps neglecta, Chaenocephalus aceratus, and a Chionodraco sp.) by means of a taxol-dependent, microtubule-affinity procedure (cf. Vallee: Journal of Cell Biology 92:435-442, 1982). MAPs from these fishes were similar to each other in electrophoretic pattern. Prominent in each preparation were proteins in the molecular weight ranges 410,000-430,000, 220,000-280,000, 140,000-155,000, 85,000-95,000, 40,000-45,000, and 32,000-34,000. The surfaces of MAP-rich microtubules were decorated by numerous filamentous projections. Exposure to elevated ionic strength released the MAPs from the microtubules and also removed the filamentous projections. Addition of fish MAPs to subcritical concentrations of fish tubulins at 0-5 degrees C induced the assembly of microtubules. Both the rate and the extent of this assembly increased with increasing concentrations of the MAPs. Sedimentation revealed that approximately six proteins, with apparent molecular weights between 60,000 and 300,000, became incorporated into the microtubule polymer. Bovine MAPs promoted microtubule formation by fish tubulin at 2-5 degrees C, and proteins corresponding to MAPs 1 and 2 co-sedimented with the polymer. MAPs from C. aceratus also enhanced the polymerization of bovine tubulin at 33 degrees C, but the microtubules depolymerized at 0 degrees C. We conclude that MAPs are part of the microtubules of Antarctic fishes, that these proteins promote microtubule assembly in much the same way as mammalian MAPs, and that they do not possess special capacities to promote microtubule assembly at low temperatures or to prevent cold-induced microtubule depolymerization.     

X-cell disease in Antarctic fishes

A number of Antarctic fish species are affected by an unusual gill condition known as X-cell disease, named in reference to morphologically similar lesions of unknown aetiology reported from northern hemisphere fishes. Despite the disease being first recorded in Antarctic fishes over 25 years ago, no progress has been made in identifying its cause or in confirming any possible relationship with northern fishes. Although once thought to be a neoplasm, observations of lesions in non-Antarctic fishes point towards a parasitic origin. The life cycle of the proposed causal organism is unknown, however, and the only stages identified are those of the eponymous cells in the lesions. Here, we show X-cells in diseased gills of the Antarctic nototheniid Trematomus bernacchii represent multinucleate cysts of an unknown parasitic organism. Furthermore, we use molecular genetic methodology to show that the organism responsible is closely related to that identified in X-cell lesions of the common European dab, Limanda limanda and that the disease thus has a global distribution. Phylogenetic tree construction based on 18S rDNA sequences confirms that X-cell organisms form a group of closely related parasites, but robust positioning of the X-cell clade in the tree awaits more extensive genetic sequencing.     

Molecular ecophysiology of Antarctic notothenioid fishes

The notothenioid fishes of the Southern Ocean surrounding Antarctica are remarkable examples of organismal adaptation to extreme cold. Their evolution since the mid-Miocene in geographical isolation and a chronically cold marine environment has resulted in extreme stenothermality of the extant species. Given the unique thermal history of the notothenioids, one may ask what traits have been gained, and conversely, what characters have been lost through change in the information content of their genomes. Two dramatic changes that epitomize such evolutionary transformations are the gain of novel antifreeze proteins, which are obligatory for survival in icy seawater, by most notothenioids and the paradoxical loss of respiratory haemoproteins and red blood cells, normally deemed indispensable for vertebrate life, by the species of a highly derived notothenioid family, the icefishes. Here, we review recent advances in our understanding of these traits and their evolution and suggest future avenues of investigation.The formerly coherent paradigm of notothenioid freeze avoidance, developed from three decades of study of antifreeze glycoprotein (AFGP) based cold adaptation, now faces challenges stemming from the recent discovery of antifreeze-deficient, yet freeze-resistant, early notothenioid life stages and from definitive evidence that the liver is not the physiological source of AFGPs in notothenioid blood. The resolution of these intriguing observations is likely to reveal new physiological traits that are unique to the notothenioids. Similarly, the model of AFGP gene evolution from a notothenioid pancreatic trypsinogen-like gene precursor is being expanded and refined based on genome-level analyses of the linked AFGP loci and their ancestral precursors. Finally, the application of comparative genomics to study evolutionary change in the AFGP genotypes of cool-temperate notothenioids from sub-Antarctic habitats, where these genes are not necessary, will contribute to the mechanistic understanding of the dynamics of AFGP gene gain and loss.In humans and most vertebrates, mutations in the alpha- or beta-globin genes or defects in globin chain synthesis are causes of severe genetic disease. Thus, the 16 species of haemoglobinless, erythrocyte-null icefishes are surprising anomalies -- in fact, they could only have evolved and thrived due to relaxed selection pressure for oxygen-binding proteins in the cold, oxygen-rich waters of the Southern Ocean. Fifteen of the sixteen icefish species have lost most of the adult alphabeta-globin locus and retain only a small 3' fragment of the alpha-globin gene. The only exception to this pattern occurs in Neopagetopsis ionah, which possesses a disrupted alphabeta-globin gene complex that probably represents a non-functional intermediate on the evolutionary pathway to near total globin gene extinction. By contrast, six of the icefish species fail to express myoglobin. The absence of myoglobin expression has occurred by several independent mutations and distinct mechanisms. Haemoprotein loss is correlated with dramatic increases in cellular mitochondrial density, heart size, blood volume and capillary bed volume. Evolution of these compensatory traits was probably facilitated by the homeostatic activity of nitric oxide, a key modulator of angiogenesis and mitochondrial biogenesis. These natural knockouts of the red blood cell lineage are an excellent genomic resource for erythroid gene discovery by comparative genomics, as illustrated for the newly described gene, bloodthirsty.     

Hepatic ultrastructural specialization in Antarctic fishes

Summary Compared with those of other vertebrate animals, the livers of Antarctic fishes have a unique type of perisinusoidal (Ito) cell. These cells were studied in 9 species with emphasis on Dissostichus mawsoni. Perisinusoidal cells are found in large numbers throughout the liver, have long cytoplasmic arms and, in Dissostichus, contain numerous lipid droplets. The extensive rough endoplasmic reticulum and prominent nucleolus are ultrastructural characteristics indicating that these cells are engaged in protein synthesis. An evolutionary specialization, perisinusoidal cells may be partially responsible for the elevated levels of protein synthesis characteristic of fishes in the Antarctic marine environment.     

南极鱼类多样性和适应性进化研究进展

南极地区是地球上唯一未被人类活动大量影响的地区, 其极端寒冷的环境为南极生物的进化提供了“温床”。过去三千万年间, 南极鱼亚目鱼类在南极海洋逐渐变冷的过程中快速进化, 从一个温暖海域的底栖祖先分化成南极海域最为多样化的鱼类类群。由于其在南极圈内和南极圈外的各种温度区间都有分布, 因而成为研究鱼类适应性进化和耐寒机制的良好生物模型。本文综述了有关南极海域鱼类区系组成与物种多样性现状, 南极鱼亚目鱼类适应低温的一系列特化的生物学性状及其关键的遗传进化机制。现有研究表明: 南极鱼类在几千万年零度以下低温环境的进化中发生了大量基因的大规模扩增和基因表达的改变, 如铁调素、卵壳蛋白和逆转座子等118个基因发生了显著的扩增。另外, 有些从南极鱼中获得的抗寒基因已经用于提高动植物低温抗性的研究并取得了良好的效果。在今后的几年中, 将会有多个南极鱼物种的全基因组得到破译, 在低温适应相关基因的功能和进化方面的研究也会更加深入, 这些研究将深入揭示低温压力下基因组的进化规律以及鱼类低温适应的分子机制。     

Feeding biomechanics of five demersal Antarctic fishes

There is scant information available on the ecomorphology of Antarctic fishes, and especially on their feeding capabilities. We measured interspecific variation in mechanical advantage (MA), force-producing capability, and suction index for the jaws of the five dominant taxa of high-Antarctic fishes: the nototheniid Trematomus bernacchii; the zoarcids Pachycara brachycephalum, Lycodichthys dearborni, and Ophthalmolycus amberensis; and the liparid Paraliparis devriesi. Analysis of variance indicated significant differences in jaw metrics, and ordinations of morphological traits identified three loosely defined groups reflecting their family-level taxonomy. Principal component analyses showed distinct segregation between the nototheniid and the liparid, indicating that they are at the extremes of the feeding performance continuum. The zoarcids fell in the middle, suggesting that they utilize a combination of feeding modes to capture prey. The liparid had the lowest MA and bite force, but a large epaxialis implied a ram-suction-feeding mode. The large adductor mandibulae in the zoarcids P. brachycephalum and L. dearborni suggest that they are capable of grasping mobile prey and manipulating sedentary, hard-shelled macroinvertebrates. The zoarcids had a smaller epaxialis than the liparid and may not be as efficient as suction-feeders. Values for mechanical advantage ratios and suction indices in Antarctic fishes were within the range known for non-Antarctic fishes. The five Antarctic species do not possess dentition specialized for durophagous feeding; however, the high mechanical advantage ratio in the nototheniid and, to a lesser extent, in the zoarcids, suggests that durophagy may be possible.     

Heat tolerance and its plasticity in Antarctic fishes

The adaptive radiation of the Antarctic notothenioid ancestral benthic fish stock within the chronic freezing waters of the Southern Ocean gave rise to five highly cold adapted families. Their stenothermy, first observed from several high-latitude McMurdo Sound species, has been of increasing recent interest given the threat of rising polar water temperatures from global climate change. In this study we determined the heat tolerance in a geographically diverse group of 11 Antarctic species as their critical thermal maximum (CTMax). When acclimatized to their natural freezing water temperatures, environmental CTMaxs ranged from 11.95 to 16.17 °C, well below those of fishes endemic to warmer waters. There was a significant regional split, with higher CTMaxs in species from the more northerly and thermally variable Seasonal Pack-ice Zone. When eight of the Antarctic species were warm acclimated to 4 °C all showed a significant increase over their environmental CTMaxs, with several showing plasticity comparable in magnitude to some far more eurythermal fishes. When the accrual of heat tolerance during acclimation was followed in three high-latitude McMurdo Sound species, it was found to develop slowly in two of them, which was correlated with their low metabolic rates.     

Disaptation and recovery in the evolution of Antarctic fishes

The radiation of notothenioid fishes provides an excellent system to explore issues of evolution and adaptation. Most studies emphasize adaptation to the extreme Antarctic environment; however, recent work provides cogent examples of disaptation or evolutionary loss of function. The nature and extent of regressive change is revealed by subsequent adaptive recovery. Ancestral notothenioids were benthic but some became secondarily pelagic through the retention of larval characters. Paedomorphosis has produced detrimental changes in lateral-line sensory systems that have been made good by compensatory adaptation. In the icefish family, compensatory adaptation has followed the loss of the oxygen-binding pigments haemoglobin and myoglobin.     

Renal glomerular evolution in Antarctic notothenioid fishes

Light and electron microscopy were used to document the degree of glomerular development in 10 species of Antarctic notothenioid fishes. When combined with results of previous studies, data revealed that 16 of 20 species inhabiting subzero sea water were aglomerular. One subantarctic and two temperate species were pauciglomerular, and an additional temperate species had a moderate number of glomeruli. Renal corpuscles were variable in number and diameter among the pauciglomerular species, and most had few patent glomerular capillaries. Radiolabelled markers indicated that the glomerular filtration rate was low in the pauciglomerular Notothenia angustata , ranging from 0.005 to 0.124 ml h⁻¹ kg⁻¹ in eight specimens. Arterial perfusion of Microfil demonstrated that arteries supplying aglomerular and pauciglomerular kidneys were confined largely to the periphery of the organ, and glomerular capillaries were absent or few in number. As ancestral notothenioids probably had glomerular kidneys, data from 20–25% of the fauna suggest that there has been an evolutionary loss of glomeruli in many species. The pattern of glomerular reduction is consistent with the hypothesis that the selective advantage of aglomerularism is in the urinary conservation of small molecular weight antifreeze glycopeptide compounds that are vital to survival in sub-zero Antarctic waters.     

Inducible heat tolerance in Antarctic notothenioid fishes

Significant increases in heat tolerance (time of survival at 14°C) were observed for some, but not all, species of notothenioid fishes collected from McMurdo Sound, Antarctica (77°51′S) following acclimation to 4°C. The increase in thermal tolerance was rapid in Trematomus bernacchii, developing within 1–2 days of acclimation to 4°C. Long-term (6–8 weeks) acclimation to 4°C led to greater heat tolerance in Trematomus pennellii than in T. bernacchii. Unlike its demersal congeners, the cryopelagic notothenioid Pagothenia borchgrevinki did not increase heat tolerance during warm acclimation. A deep-living zoarcid fish, Lycodichthys dearborni, also failed to increase heat tolerance, but survived significantly (> threefold) longer at 14°C than the notothenioids.     

Comparison of antifreeze glycopeptides from several Antarctic fishes

Summary Antifreeze glycopeptides have been isolated from eight species of antarctic fish representing the families Nototheniidae, Channichthyidae, and Bathydraconidae. Amino acid analysis indicates that the glycopeptides have amino acid compositions identical to previously studied antarctic nototheniids. The sizes of the glycopeptides show some heterogeneity although they all span the molecular weight range of approximately 34,000 to 2,500 daltons. The species studied here are representative of most of the ecological habitats of the Antarctic Ocean.