Histological evaluation of the distribution of systemic AA-amyloidosis in nine domestic shorthair cats

Amyloidosis is a group of protein-misfolding disorders characterized by the accumulation of amyloid in organs, both in humans and animals. AA-amyloidosis is considered a reactive type of amyloidosis and in humans is characterized by the deposition of AA-amyloid fibrils in one or more organs. In domestic shorthair cats, AA-amyloidosis was recently reported to be frequent in shelters. To better characterize this pathology, we report the distribution of amyloid deposits and associated histological lesions in the organs of shelter cats with systemic AA-amyloidosis. AA-amyloid deposits were identified with Congo Red staining and immunofluorescence. AA-amyloid deposits were then described and scored, and associated histological lesions were reported. Based on Congo Red staining and immunofluorescence nine shelter cats presented systemic AA-amyloidosis. The kidney (9/9), the spleen (8/8), the adrenal glands (8/8), the small intestine (7/7) and the liver (8/9) were the organs most involved by amyloid deposits, with multifocal to diffuse and from moderate to severe deposits, both in the organ parenchyma and/or in the vascular compartment. The lung (2/9) and the skin (1/8) were the least frequently involved organs and deposits were mainly focal to multifocal, mild, vascular and perivascular. Interestingly, among the organs with fibril deposition, the stomach (7/9), the gallbladder (6/6), the urinary bladder (3/9), and the heart (6/7) were reported for the first time in cats. All eye, brain and skeletal muscle samples had no amyloid deposits. An inflammatory condition was identified in 8/9 cats, with chronic enteritis and chronic nephritis being the most common. Except for secondary cell compression, other lesions were not associated to amyloid deposits. To conclude, this study gives new insights into the distribution of AA-amyloid deposits in cats. A concurrent chronic inflammation was present in almost all cases, possibly suggesting a relationship with AA-amyloidosis

Histological evaluation of the distribution of systemic AA-amyloidosis in nine domestic shorthair cats.

Amyloidosis is a group of protein-misfolding disorders characterized by the accumulation of amyloid in organs, both in humans and animals. AA-amyloidosis is considered a reactive type of amyloidosis and in humans is characterized by the deposition of AA-amyloid fibrils in one or more organs. In domestic shorthair cats, AA-amyloidosis was recently reported to be frequent in shelters. To better characterize this pathology, we report the distribution of amyloid deposits and associated histological lesions in the organs of shelter cats with systemic AA-amyloidosis. AA-amyloid deposits were identified with Congo Red staining and immunofluorescence. AA-amyloid deposits were then described and scored, and associated histological lesions were reported. Based on Congo Red staining and immunofluorescence nine shelter cats presented systemic AA-amyloidosis. The kidney (9/9), the spleen (8/8), the adrenal glands (8/8), the small intestine (7/7) and the liver (8/9) were the organs most involved by amyloid deposits, with multifocal to diffuse and from moderate to severe deposits, both in the organ parenchyma and/or in the vascular compartment. The lung (2/9) and the skin (1/8) were the least frequently involved organs and deposits were mainly focal to multifocal, mild, vascular and perivascular. Interestingly, among the organs with fibril deposition, the stomach (7/9), the gallbladder (6/6), the urinary bladder (3/9), and the heart (6/7) were reported for the first time in cats. All eye, brain and skeletal muscle samples had no amyloid deposits. An inflammatory condition was identified in 8/9 cats, with chronic enteritis and chronic nephritis being the most common. Except for secondary cell compression, other lesions were not associated to amyloid deposits. To conclude, this study gives new insights into the distribution of AA-amyloid deposits in cats. A concurrent chronic inflammation was present in almost all cases, possibly suggesting a relationship with AA-amyloidosis

Glycosylation tunes neuroserpin physiological and pathological properties

Neuroserpin (NS) is a member of the serine protease inhibitors superfamily. Specific point mutations are responsible for its accumulation in the endoplasmic reticulum of neurons that leads to a pathological condition named familial encephalopathy with neuroserpin inclusion bodies (FENIB). Wild-type NS presents two N-glycosylation chains and does not form polymers in vivo, while non-glycosylated NS causes aberrant polymer accumulation in cell models. To date, all in vitro studies have been conducted on bacterially expressed NS, de facto neglecting the role of glycosylation in the biochemical properties of NS. Here, we report the expression and purification of human glycosylated NS (gNS) using a novel eukaryotic expression system, LEXSY. Our results confirm the correct N-glycosylation of wild-type gNS. The fold and stability of gNS are not altered compared to bacterially expressed NS, as demonstrated by the circular dichroism and intrinsic tryptophan fluorescence assays. Intriguingly, gNS displays a remarkably reduced polymerisation propensity compared to non-glycosylated NS, in keeping with what was previously observed for wild-type NS in vivo and in cell models. Thus, our results support the relevance of gNS as a new in vitro tool to study the molecular bases of FENIB

physical activity monitoring devices energy expenditure comparison in a setting of free living activities

The aim of this study was to evaluate the validity of Energy Expenditure (EE) estimation provided by 3 wearable devices [Fitbit-One (FO), Sensewear Armband (AR) and Actiheart (AC)] in a setting of free-living activities. 43 participants (24 females; 23.4±.4,5yrs) performed 9 activities: sedentary (watching video, reading), walking (on treadmill and outdoor), running (on treadmill and outdoor) and moderate-to-vigorous activities (Wii gaming, taking the stairs and playing football). Mean Absolute Percentage Error (MAPE) and Pearson's correlation were calculated to assess the validity of each instrument in comparison to a portable metabolic analyser (PMA). In overall comparison MAPE's were 7,7% for AR (r=.86; p<.0001), 8,6% for FO (r=.69; P<.001), and 11.6% for AC (r=.81; p<.0001). These findings support the accuracy of the wearables. The AR was the most accurate in the whole protocol. However, MAPE results suggest that devices algorithms should be improved for better measure of EE during moderate-to-vigorous activities

Renal amyloid‐A amyloidosis in cats: Characterization of proteinuria and biomarker discovery, and associations with kidney histology

BackgroundAmyloid A (AA) amyloidosis is a protein misfolding disease arising from serum amyloid A (SAA). Systemic AA amyloidosis recently was shown to have a high prevalence in shelter cats in Italy and was associated with azotemia and proteinuria.ObjectivesInvestigate urine protein profiles and diagnostic biomarkers in cats with renal AA amyloidosis.AnimalsTwenty‐nine shelter cats.MethodsCase‐control study. Cats with renal proteinuria that died or were euthanized between 2018 and 2021 with available necropsy kidney, liver and spleen samples, and with surplus urine collected within 30 days before death, were included. Histology was used to characterize renal damage and amyloid amount and distribution; immunohistochemistry was used to confirm AA amyloidosis. Urine protein‐to‐creatinine (UPC) and urine amyloid A‐to‐creatinine (UAAC) ratios were calculated, and sodium dodecyl sulfate‐agarose gel electrophoresis (SDS‐AGE) and liquid chromatography‐mass spectrometry (LC‐MS) of proteins were performed.ResultsTwenty‐nine cats were included. Nineteen had AA amyloidosis with renal involvement. Cats with AA amyloidosis had a higher UPC (median, 3.9; range, 0.6‐12.7 vs 1.5; 0.6‐3.1; P = .03) and UAAC ratios (median, 7.18 × 10$^{−3}$; range, 23 × 10$^{−3}$‐21.29 × 10$^{−3}$ vs 1.26 × 10$^{−3}$; 0.21 × 10$^{−3}$‐6.33 × 10$^{−3}$; P = .04) than unaffected cats. The SDS‐AGE identified mixed‐type proteinuria in 89.4% of cats with AA amyloidosis and in 55.6% without AA amyloidosis (P = .57). The LC‐MS identified 63 potential biomarkers associated with AA amyloidosis (P < .05). Among these, urine apolipoprotein C‐III was higher in cats with AA amyloidosis (median, 1.38 × 10$^{7}$; range, 1.85 × 10$^{5}$‐5.29 × 10$^{7}$ vs 1.76 × 10$^{6}$; 0.0 × 10$^{0}$‐1.38 × 10$^{7}$; P = .01). In the kidney, AA‐amyloidosis was associated with glomerulosclerosis (P = .02) and interstitial fibrosis (P = .05).Conclusions and Clinical ImportanceRenal AA amyloidosis is associated with kidney lesions, increased proteinuria and increased urine excretion of SAA in shelter cats. Additional studies are needed to characterize the role of lipid transport proteins in the urine of affected cats

Cryo-EM structure of ex vivo fibrils associated with extreme AA amyloidosis prevalence in a cat shelter

AA amyloidosis is a systemic disease characterized by deposition of misfolded serum amyloid A protein (SAA) into cross-β amyloid in multiple organs in humans and animals. AA amyloidosis occurs at high SAA serum levels during chronic inflammation. Prion-like transmission was reported as possible cause of extreme AA amyloidosis prevalence in captive animals, e.g. 70% in cheetah and 57–73% in domestic short hair (DSH) cats kept in zoos and shelters, respectively. Herein, we present the 3.3 Å cryo-EM structure of AA amyloid extracted post-mortem from the kidney of a DSH cat with renal failure, deceased in a shelter with extreme disease prevalence. The structure reveals a cross-β architecture assembled from two 76-residue long proto-filaments. Despite >70% sequence homology to mouse and human SAA, the cat SAA variant adopts a distinct amyloid fold. Inclusion of an eight-residue insert unique to feline SAA contributes to increased amyloid stability. The presented feline AA amyloid structure is fully compatible with the 99% identical amino acid sequence of amyloid fragments of captive cheetah

Cryo-EM structure of ex vivo fibrils associated with extreme AA amyloidosis prevalence in a cat shelter.

AA amyloidosis is a systemic disease characterized by deposition of misfolded serum amyloid A protein (SAA) into cross-β amyloid in multiple organs in humans and animals. AA amyloidosis occurs at high SAA serum levels during chronic inflammation. Prion-like transmission was reported as possible cause of extreme AA amyloidosis prevalence in captive animals, e.g. 70% in cheetah and 57-73% in domestic short hair (DSH) cats kept in zoos and shelters, respectively. Herein, we present the 3.3 Å cryo-EM structure of AA amyloid extracted post-mortem from the kidney of a DSH cat with renal failure, deceased in a shelter with extreme disease prevalence. The structure reveals a cross-β architecture assembled from two 76-residue long proto-filaments. Despite >70% sequence homology to mouse and human SAA, the cat SAA variant adopts a distinct amyloid fold. Inclusion of an eight-residue insert unique to feline SAA contributes to increased amyloid stability. The presented feline AA amyloid structure is fully compatible with the 99% identical amino acid sequence of amyloid fragments of captive cheetah

AA-amyloidosis in cats (Felis catus) housed in shelters.

Systemic AA-amyloidosis is a protein-misfolding disease characterized by fibril deposition of serum amyloid-A protein (SAA) in several organs in humans and many animal species. Fibril deposits originate from abnormally high serum levels of SAA during chronic inflammation. A high prevalence of AA-amyloidosis has been reported in captive cheetahs and a horizontal transmission has been proposed. In domestic cats, AA-amyloidosis has been mainly described in predisposed breeds but only rarely reported in domestic short-hair cats. Aims of the study were to determine AA-amyloidosis prevalence in dead shelter cats. Liver, kidney, spleen and bile were collected at death in cats from 3 shelters. AA-amyloidosis was scored. Shedding of amyloid fibrils was investigated with western blot in bile and scored. Descriptive statistics were calculated. In the three shelters investigated, prevalence of AA-amyloidosis was 57.1% (16/28 cats), 73.0% (19/26) and 52.0% (13/25), respectively. In 72.9% of cats (35 in total) three organs were affected concurrently. Histopathology and immunofluorescence of post-mortem extracted deposits identified SAA as the major protein source. The duration of stay in the shelters was positively associated with a histological score of AA-amyloidosis (B = 0.026, CI95% = 0.007-0.046; p = 0.010). AA-amyloidosis was very frequent in shelter cats. Presence of SAA fragments in bile secretions raises the possibility of fecal-oral transmission of the disease. In conclusion, AA-amyloidosis was very frequent in shelter cats and those staying longer had more deposits. The cat may represent a natural model of AA-amyloidosis

The two tryptophans of β2-microglobulin have distinct roles in function and folding and might represent two independent responses to evolutionary pressure

We have recently discovered that the two tryptophans of human β2-microglobulin have distinctive roles within the structure and function of the protein. Deeply buried in the core, Trp95 is essential for folding stability, whereas Trp60, which is solvent-exposed, plays a crucial role in promoting the binding of β2-microglobulin to the heavy chain of the class I major histocompatibility complex (MHCI). We have previously shown that the thermodynamic disadvantage of having Trp60 exposed on the surface is counter-balanced by the perfect fit between it and a cavity within the MHCI heavy chain that contributes significantly to the functional stabilization of the MHCI. Therefore, based on the peculiar differences of the two tryptophans, we have analysed the evolution of β2-microglobulin with respect to these residues