The role of lysosomes and autophagosomes in frontotemporal lobar degeneration.

INTRODUCTION: Cell biological and genetic evidence implicate failures in degrading aggregating proteins, such as tau and TDP-43, through the autophagy or lysosomal pathways in the pathogenesis of frontotemporal lobar degeneration (FTLD). METHODS: We investigated changes in the degradative pathways in 60 patients with different pathological or genetic forms of FTLD employing immunohistochemistry for marker proteins such as lysosomal-associated membrane proteins 1 (LAMP-1) and 2 (LAMP-2), cathepsin D (CTSD) and microtubule-associated protein 1 light chain 3 alpha (LC3A). Immunostained sections were qualitatively and semi-quantitatively assessed for the appearance, distribution and intensity of staining in neurones of the dentate gyrus (DG) and CA4 region of the hippocampus, and the temporal cortex (Tcx). RESULTS: Lower levels of neuronal LAMP-1 immunostaining were present in the DG and Tcx in FTLD-tau compared to FTLD-TDP. There was less LAMP-1 immunostaining in FTLD-tau with MAPT mutations, and FTLD-tau with Pick bodies, compared to FTLD-TDP types A and B, and less LAMP-1 immunostaining in FTLD-TDP type C than in FTLD-TDP types A and B. There was greater LAMP-1 immunostaining in GRN mutation which may reflect the underlying type A histology rather than mutation. There were no differences in neuronal LAMP-2, CTSD, EEA-1 or LC3A immunostaining between any of the five FTLD histological or four genetic groups, nor between FTLD-TDP and FTLD-tau. CONCLUSIONS: The underlying pathological mechanism in FTLD-tau may lie with a relative deficiency of lysosomes, or defective vesicular transport, whereas the failure to clear TDP-43 aggregates may lie with lysosomal dysfunction rather than a lack of available lysosomes or degradative enzymes.

Patterns of cerebral amyloid angiopathy define histopathological phenotypes in Alzheimer's disease.

AIMS: Pathological heterogeneity of Aß deposition in senile plaques (SP) and cerebral amyloid angiopathy (CAA) in Alzheimer's disease (AD) has been long noted. The aim of this study was to classify cases of AD according to their pattern of Aß deposition, and to seek factors which might predict, or predispose towards, this heterogeneity. METHODS: The form, distribution and severity of Aß deposition (as SP and/or CAA) was assessed semiquantitatively in immunostained sections of frontal, temporal and occipital cortex from 134 pathologically confirmed cases of AD. RESULTS: Four patterns of Aß deposition were defined. Type 1 describes cases predominantly with SP, with or without CAA within leptomeningeal vessels alone. Type 2 describes cases where, along with many SP, CAA is present in both leptomeningeal and deeper penetrating arteries. Type 3 describes cases where capillary CAA is present along with SP and arterial CAA. Type 4 describes a predominantly vascular phenotype, where Aß deposition is much more prevalent in and around blood vessels, than as SP. As would be anticipated from the group definitions, there were significant differences in the distribution and degree of CAA across the phenotype groups, although Aß deposition as SP did not vary. There were no significant differences between phenotype groups with regard to age of onset, age at death, disease duration and brain weight, or disease presentation. Women were over-represented in the type 1 phenotype and men in type 2. Genetically, type 3 (capillary subtype) cases were strongly associated with possession of the APOE ε4 allele. CONCLUSIONS: This study offers an alternative method of pathologically classifying cases of AD. Further studies may derive additional genetic, environmental or clinical factors which associate with, or may be responsible for, these varying pathological presentations of AD.

Nuclear carrier and RNA-binding proteins in frontotemporal lobar degeneration associated with fused in sarcoma (FUS) pathological changes.

AIMS: We aimed to investigate the role of the nuclear carrier and binding proteins, transportin 1 (TRN1) and transportin 2 (TRN2), TATA-binding protein-associated factor 15 (TAF15) and Ewing's sarcoma protein (EWS) in inclusion body formation in cases of frontotemporal lobar degeneration (FTLD) associated with fused in sarcoma protein (FTLD-FUS). METHODS: Eight cases of FTLD-FUS (five cases of atypical FTLD-U, two of neuronal intermediate filament inclusion body disease and one of basophilic inclusion body disease) were immunostained for FUS, TRN1, TRN2, TAF15 and EWS. Ten cases of FTLD associated with TDP-43 inclusions served as reference cases. RESULTS: The inclusion bodies in FTLD-FUS contained TRN1 and TAF15 and, to a lesser extent, EWS, but not TRN2. The patterns of immunostaining for TRN1 and TAF15 were very similar to that of FUS. None of these proteins was associated with tau or TDP-43 aggregations in FTLD. CONCLUSIONS: Data suggest that FUS, TRN1 and TAF15 may participate in a functional pathway in an interdependent way, and imply that the function of TDP-43 may not necessarily be in parallel with, or complementary to, that of FUS, despite each protein sharing many similar structural elements.

The effect of aging on skeletal muscle capillarization in a murine model.

Capillarization of skeletal muscle has been reported to be both maintained and reduced with advancing age. This conflict may represent methodological differences between biopsy studies. We have examined capillarization throughout two muscles, soleus and extensor digitorum longus (EDL), from a well-established colony of aging mice, and related this to fiber number (C/F ratio) and type. Labeling of muscle capillaries was performed with the biotinylated Griffonia (Bandeiraea) simplicifolia lectin (GSL 1) using immunochemistry. The results showed a significant increase in the C/F ratio in the aged mice when compared with the younger (6-month mice soleus = 1.296, 95% CI 1.226-1.366 vs 28-month mice soleus = 1.530, 95% CI 1.488-1.572, p <.001; 6-month mice EDL = 0.881, 95% CI 0.751-1.011 vs 28-month mice EDL = 1.124, 95% CI 1.028-1.220, p = .017). These differences could not be accounted for by changes in fiber type but may reflect loss of fibers. Alternatively, there may be increased angiogenic drive or a failure of downregulation of angiogenesis.

Age-related postreceptor mechanisms: changes in adenylate cyclase but not phosphodiesterase in isolated mouse renal medullary collecting ducts.

Urine concentrating ability declines with increasing age, partly due to an impaired response of kidney medullary collecting ducts to the antidiuretic hormone, vasopressin. We investigated this change in isolated mouse medullary collecting ducts by measuring the activity of adenylate cyclase and phosphodiesterase, which catalyse the formation and hydrolysis of cAMP, respectively. Adenylate cyclase activity was measured in the presence of vasopressin (which stimulates adenylate cyclase via the receptor) or forskolin (which directly stimulates the catalytic subunit). We showed an age-related decrease in the catalytic subunit of adenylate cyclase, and no difference in the activity of phosphodiesterase, indicating that a reduction in the catalytic subunit of adenylate cyclase contributes towards the age-related decrease in cyclic AMP response of kidney to vasopressin.

Age-associated changes in neuroaxonal transport in the hypothalamo-neurohypophysial system of the mouse.

In this study, we investigated age-associated changes in neuroaxonal transport of the hormone vasopressin (AVP) and its associated neurophysin (NPII), from the supraoptic nucleus (SON) of the hypothalamus to the neurohypophysis. C57BL/Icrfat male mice of 6 and 28 months of age were injected in the hypothalamus with L-[35S]cysteine. Animals were killed up to 2.25 h after injection and NPII and AVP from the SON and neurohypophysis were separated using HPLC, and the fractions counted for radioactivity. In the SON, radiolabelled NPII and AVP were first detected after 0.50 h in both young and old mice. There was no significant difference between the age-groups in the incorporation of radiolabel over the time course studied. Radiolabelled NPII in the neurohypophysis was significantly above background after 1.25 h in the young, and after 1.50 h in the old mice. The differences between the two age groups was significant (P = 0.05). Radiolabelled AVP followed a similar trend, but was not significantly above background until 1.50 h in the young and 1.75 h in the old. The differences between the two age groups was on the point of significance (P = 0.056). These results indicate a significant reduction of up to 25% in the rate of axonal transport of neurohypophyseal peptides with advancing age.

The ageing hypothalamo-neurohypophysial system. An analysis of the neurohypophysis in normal hydration, osmotic loading and rehydration.

Age-associated changes in the structure and function of the neurohypophysis may contribute to the decreased ability to conserve water in older animals. We investigated the neurohypophyses of 6 and 28-month-old male mice using radioimmunoassay and quantitative morphological techniques. The dry-weight and volume of the neurohypophysis increased significantly with age but the quantity of vasopressin in the gland remained constant. Oxytocin levels decreased with age. A quantitative morphological analysis was performed on the compartments of the neurohypophysis from male mice of 6 and 28 months of age which were either normally hydrated, osmotically loaded, or osmotically loaded and rehydrated. The absolute volumes of the axon endings, swellings, their constituent organelles and the axon terminals containing degenerating subcellular components were determined. The design of the analysis allowed us to examine both age-related changes and statistical interactions between the age of the animal and the behaviour of a variable during the osmotic loading/rehydration phase of the experiment. There was a significant age-related reduction in the volume of the neurohypophysis occupied by the endings and swellings. The diameters of the neurosecretory granules found in the endings were significantly smaller than those in the swellings in both age groups but the size difference was greater in the young animals. Dehydration and subsequent rehydration of old male mice leads to extensive re-modelling of the neurohypophysial compartments and subcellular organelles to the configuration found in the adult animal.

Renal vasopressin receptors in ageing C57BL/Icrfat mice.

The mechanism of water conservation is impaired in ageing mammals. An age-related defect in the release of vasopressin has been implicated but, more recently, attention has moved to the renal component of the water conservation mechanism. Previous studies using renal cells prepared from mice of different ages have shown that the threshold dose of vasopressin required to elicit a significant rise in cyclic AMP (cAMP) was greater in older animals. The dose-response curve was moved to the right in 35-month-old mice, i.e. the concentration of vasopressin required to give maximum cAMP output was increased. To investigate this further we examined the binding of vasopressin to renal medullary cells maintained in short-term culture, to determine whether the decreased response of cAMP levels to vasopressin is due to changes in hormone-receptor interaction. In 6-month-old male mice the dissociation constant (Kd) was 2.38 nmol/l and the maximum binding of the hormone (Bmax) was 47.6 fmol/10(6) cells, and at 30 months of age Kd was 2.37 nmol/l and Bmax was 47.0 fmol/10(6) cells. In female mice the changes were more complicated because the data for the 6-month-old mice could be split into two groups. It is concluded that there are no age-related differences in the numbers of receptors or their affinity for vasopressin, and that the decreased cAMP response is probably associated with post-receptor mechanisms in this species.

Effect of ageing on cyclic AMP output by renal medullary cells in response to arginine vasopressin in vitro in C57BL/Icrfat mice.

The effect of age on the cyclic AMP (cAMP) response to increases in the concentration of arginine vasopressin in the presence of isobutyl methylxanthine (100 mumol/l) was studied in an in-vitro renal cell suspension prepared from C57BL/Icrfat mice at 6, 12, 18, 24, 29 and 35 months of age. Comparison of the response of the preparation to vasopressin, calcitonin and parathyroid hormone suggested that it was enriched with renal medullary cells. Basal cAMP output was similar throughout but the threshold dose of vasopressin increased from 1 X 10(-11) mol/l (6, 12 and 18 months of age) to 1 X 10(-10) mol/l (24, 29 and 35 months of age). The dose-response curve in 35-month-old mice was shifted to the right with the concentration of vasopressin required to give half maximal cAMP increased from 9.4 +/- 0.37 X 10(-11) mol/l (6 months) to 3.5 +/- 1.6 X 10(-10) mol/l (35 months). Maximum cAMP output at 1 X 10(-9) mol/l was also reduced in the same animals (stimulated:basal ratio, 51.22 +/- 19.12 at 6 months; 11.50 +/- 6.02 at 35 months). The results suggest that the lack of renal response to vasopressin in terms of cAMP metabolism may play a role in the well-documented age-related decline in urine-concentrating ability in experimental animals and elderly people.