The human embryonic-fetal kidney endoplasmic reticulum phosphate-pyrophosphate transport protein

Glucose-6-phosphatase is a multicomponent endoplasmic reticulum system comprising at least six different proteins, including a lumenal enzyme and several transport proteins. One of the transport proteins, T₂, transports the substrate pyrophosphate and the product phosphate and its genetic deficiency is termed type 1c glycogen storage disease. We have used anti-T₂ antibodies for immunohistochemistry with image analysis and kinetic analysis of the glucose-6-phosphatase system to study for the temporal and spatial development of T₂ in human embryonic and fetal kidney. In metanephric kidney, there is an early predominance of T₂ expression in the ureteric bud derivatives and this changes with ontogeny such that developing nephrons, particularly proximal tubules, become dominant by mid-gestation. T₂ has the same spatial and temporal pattern as the glucose-6-phosphatase enzyme in both mesonephric and metanephric kidney. Pyrophosphate transport capacity is appropriate for the amount of glucose-6-phosphatase activity present in mid-gestation fetal kidney, in contrast to liver, where pyrophosphate transport capacity is developmentally delayed. Increasing knowledge of the temporal and spatial expression of the glucose-6-phosphatase proteins and their catalytic roles in early human development is essential for the elucidation of the aetiology of renal disease in both type I glycogen storage diseases and the developmental disorders of the glucose-6-phosphatase system.     

Neuronal involution during ageing. Ultrastructural study in the rat cerebellum.

Involutive phenomena have been investigated by electron microscopy in the Purkinje Pk neuron of the cerebellar cortex of the aging rat. The still limited number of specimens available to date, however, suggest an age-related progression of morphological and functional deteriorations involving particularly the intraneuronal "nucleus-ribosome system" (NRS). The impairments are characterized by changes in the nucleolar texture. These alterations are accompanied by modifications in the repartition and relative proportion of RNP components of the nucleolus. In addition, other nuclear elements such as interchromatin and perichromatin granules may vary in importance with age. Recognizable changes in the ribosomal constituents of the NRS are evidenced by modifications in the density and distribution of free ribosomes. An altered structure and organization of GER cisternae are also evident. Furthermore, "light" cytoplasmic areas, an increased evidence of neurotubules and the gradual congestion of the pericaryon by age pigments are other valuable ultrastructural features that may be regarded as part of the sequence of morphologic events occurring during neuonal ageing. The above ultrastructural data will subsequently form the basis of a model of ageing in the nerve cell, which will complete the previously proposed model of neuronal maturation. Therefore, this long-term study essentially purports the investigation of subcellular events taking place in the Pk neuron all along the normal life span in rats. This model will also be used to evaluate the changes in the sequence and the reinforcement of the processes of evolution versus involution as affected by certain xenobiotics, such as abused drugs(alcohol and narcotics). The intraneuronal modifications found in the nuclear and cytoplasmic structures of the NRS could possibly reflect the molecular dysfunction related to the production of various types of RNA and neuronal proteins. This hypothesis is supported by biochemical data obtained from analysis of the brain of aged animals. Ultrastructural and biochemical data appear to be in good agreement with the neurophysiologic interpretation of a slow-down and reduced efficiency of the CNS during the progressive development of senescence in human and animal subjects.     

Caffeine-induced calcium release from isolated sarcoplasmic reticulum of rabbit skeletal muscle

The essential conditions for the Ca²⁺ releasing action of caffeine from isolated sarcoplasmic reticulum (SR) of rabbits were evaluated by an investigation into the effects of Ca²⁺, Mg²⁺, MgATP^2–, and ATP concentration, ionic strength, and degree of loading. The heavy fraction (4,500×g) of the reticulum was used. Except for the study on degree of loading, 0.2 mg protein·ml^–1 SR was loaded actively with 0.02 mM⁴⁵CaCl₂, resulting in >90 nmol·mg protein^–1 at steady state, and then the effects of various parameters with or without (control) caffeine were tested.It was found that (1) caffeine induces a transient, dosedependent release of Ca²⁺, (2) the absolute amount of Ca²⁺ released by caffeine increases with the Ca²⁺ load of the SR, (3) increasing the ionic strength () from 0.09 to 0.3 lowers the threshold concentration of caffeine, (4) the SR is refractory to a repeated challenge by a caffeine concentration causing maximal effect, (5) caffeine-induced Ca²⁺ release increases with increasing (a) external Ca²⁺ concentrations up to 5 M total Ca²⁺ (or 3 M free Ca²⁺) and (b) free ATP concentrations up to 0.45 mM, and (6) caffeine-induced Ca²⁺ release is not affected by changes of either the Mg²⁺ or the MgATP^2– concentration.     

线粒体自噬与骨髓间充质干细胞成软骨分化的关联

背景:已有研究发现线粒体自噬在软骨缺损修复过程中发挥重要作用,因此有必要研究线粒体自噬在骨髓间充质干细胞成软骨分化过程中的作用及其调控方法,为进一步阐明软骨诱导机制并为理解生物材料如何调控骨髓间充质干细胞分化命运奠定基础。目的:建立并验证线粒体状态和自噬程度的表征方法,观察线粒体自噬与软骨细胞发育、骨髓间充质干细胞成软骨分化的关系。方法:分离培养新生乳兔、1月龄兔、18月龄兔软骨细胞和新生乳兔骨髓间充质干细胞,按照试剂盒说明方法进行线粒体标记Mito Tracker Red染色。选取第2代骨髓间充质干细胞,用完全软骨诱导培养基培养,在培养第1,3,7天按照试剂盒说明方法进行线粒体标记Mito Tracker Red染色。选取第2代骨髓间充质干细胞,用完全软骨诱导培养基培养24 h后,诱导组加入10μmol/L自噬诱导剂雷帕霉素干预10 h,抑制组加入5μmol/L自噬抑制剂氯喹干预10 h,此后换用完全软骨培养基培养,在培养第1,3,7天用线粒体自噬试剂盒Mitophagy Detection Kit染色并观察线粒体自噬情况。结果与结论:①不同兔龄软骨细胞线粒体染色:低兔龄软骨细胞中线粒体数目较高兔龄软骨细胞多;②骨髓间充质干细胞和幼兔软骨细胞线粒体染色:软骨细胞的线粒体形态呈点状,而骨髓间充质干细胞的线粒体形态呈线状;③骨髓间充质干细胞分化过程中线粒体染色:随诱导培养天数的增加,骨髓间充质干细胞中线粒体的形态由最初的网状变为点状,骨髓间充质干细胞骨架中促进自噬形成的微丝结构也逐渐减少;④线粒体自噬对成软骨分化的影响:雷帕霉素促进了线粒体自噬途径,进而促进骨髓间充质干细胞的成软骨分化。     

Ultrastructural characteristics of the interdigitating dendritic cell in dermatopathic lymphadenopathy of mycosis fungoides patients

The ultrastructure of interdigitating dendritic cells in dermatopathic lymphadenopathy from patients with mycosis fungoides was investigated. The most remarkable findings were: a marked hypertrophy of the smooth endoplasmic reticulum with a peculiar concentric arrangements of the cisternae around lipid droplets, an abundance of cytoplasmic lipid droplets and the presence of paranuclear chromatoid-like bodies. It is suggested that these ultrastructural features may correspond to some unknown functional capability of the interdigitating dendritic cell.     

Unusual familial cardiomyopathy with storage of intermediate filaments in the cardiac muscular cells

Summary Unusual histological and ultrastructural changes in cardiac muscle cells have been found in 3 brothers with progressive myocardial deficiency. Histologically, this cardiomyopathy was characterized by massive storage of PAS-negative proteinaceous material in most cardiac muscle cells. The electron microscope showed that this material consisted of sinuous filaments, 7–10 nm in diameter, similar to the intermediate filaments normally present in cardiac muscle cells. Filament storage coincided with the disintegration of neighbouring myofibrils, with particular change in Z bands giving rise to rod-like bodies and more complex structures formed by the association of Z band material and sarcoplasmic reticulum (SR) tubules. Filament storage and myofibrillar disintegration always occurred in areas where the SR developed and involuted extensively. Relatively high glycogen accumulation also occurred, in close relation to the SR changes. Discrete SR proliferation, glycogen overload and filament deposits were observed in a few skeletal fibres.These observations suggest that disturbance in the metabolism of desmin (protein subunit of intermediate filaments and a fundamental component of Z bands) might be involved in this type of cardiomyopathy. The influence of a chronic defect in calcium regulation might also be envisaged in view of the marked SR abnormalities.     

Antigen presentation in SARS-CoV-2 infection: the role of class I HLA and ERAP polymorphisms

Given the highly polymorphic nature of Human Leukocyte Antigen (HLA) molecules, it is not surprising that they function as key regulators of the host immune response to almost all invading pathogens, including SARS-CoV-2, the etiological agent responsible for the recent COVID-19 pandemic. Several correlations have already been established between the expression of a specific HLA allele/haplotype and susceptibility/progression of SARS-CoV-2 infection and new ones are continuously emerging. Protective and harmful HLA variants have been described in both mild and severe forms of the disease, but considering the huge amount of existing variants, the data gathered in such a brief span of time are to some extent confusing and contradictory. The aim of this mini-review is to provide a snap-shot of the main findings so far collected on the HLA-SARS-CoV-2 interaction, so as to partially untangle this intricate yarn. As key factors in the generation of antigenic peptides to be presented by HLA molecules, ERAP1 and ERAP2 role in SARS-CoV-2 infection will be revised as well.     

Effects of enflurane on excitation-contraction coupling in frog skeletal muscle fibers

The effect of enflurane on the excitation-contraction (E-C) coupling of frog skeletal muscle fiber was studied.Low (19.13±0.70 mg%) and high (108.52±4.52 mg%) concentrations of enflurane did not cause substantial changes of the resting membrane potential but a high concentration of enflurane reduced the peak of action potentials and prolonged the duration measured at 50% peak amplitude.During the twitch response, enflurane reduced the peak response of light signal measured with calcium-sensitive photoprotein, aequorin, butpeak tension was not diminished by a low concentration of enflurane. A high concentration of enflurane remarkably inhibited both light signal and tension.During tetanus, a low concentration of enflurane partially abolished light signal and tension. At the high concentration, this inhibitory effect was pronuunced and action potentials were only observed in the initial phase of tetanic stimulation.Enflurane enhanced the increase of light signal observed in rapid cooling contracture.In glycerinated muscle fiber, enflurane shifted the pCa-tension relation to the left in low calcium and suppressed maximally activated tension at high calcium concentration.Inhibitory effect of enflurane on light signal in twitch response without abolition of action potential suggests that enflurane might inhibit E-C coupling in frog skeletal muscle fiber (but it enhances direct calcium release from SR induced by caffeine) and increase calcium sensitivity of contractile element.     

Effect of chloride on Ca2+ release from the sarcoplasmic reticulum of mechanically skinned skeletal muscle fibres

 The effect of intracellular Cl^– on Ca²⁺ release in mechanically skinned fibres of rat extensor digitorum longus (EDL) and toad iliofibularis muscles was examined under physiological conditions of myoplasmic [Mg²⁺] and [ATP] and sarcoplasmic reticulum (SR) Ca²⁺ loading. Both in rat and toad fibres, the presence of 20 mM Cl^–in the myoplasm increased Ca²⁺ leakage from the SR at pCa (i.e. –log₁₀ [Ca²⁺]) 6.7, but not at pCa 8. Ca²⁺ uptake was not significantly affected by the presence of Cl^–. This Ca²⁺-dependent effect of Cl^– on Ca²⁺ leakage was most likely due to a direct action on the ryanodine receptor/Ca²⁺ release channel, and could influence channel sensitivity and the resting [Ca²⁺] in muscle fibres in vivo. In contrast to this effect, acute addition of 20 mM Cl^– to the myoplasm caused a 40–50% reduction in Ca²⁺ release in response to a low caffeine concentration both in toad and rat fibres. One possible explanation for this latter effect is that the addition of Cl^– induces a potential across the SR (lumen negative) which might reduce Ca²⁺ release via several different mechanisms. Received: 20 October 1997 / Received after revision: 1 December 1997 / Accepted: 2 December 1997