New clinical forms of hereditary apoA-I amyloidosis entail both glomerular and retinal amyloidosis.

Apolipoprotein A1 amyloidosis (ApoAI) results from specific mutations in the APOA1 gene causing abnormal accumulation of amyloid fibrils in diverse tissues. The kidney is a prominent target tissue in ApoAI amyloidosis with a remarkable selectivity for the renal medulla. Here, we investigated six French families with ApoAI Glu34Lys, p.His179Profs∗47, and a novel p.Thr185Alafs∗41 variant revealing unprecedented clinical association of a glomerular with a retinal disease. Comprehensive clinicopathological, molecular and proteomics studies of numerous affected tissues ensured the correlation between clinical manifestations, including novel unrecognized phenotypes, and apoA-I amyloid deposition. These ophthalmic manifestations stemmed from apoA-I amyloid deposition, highlighting that the retina is a previously unrecognized tissue affected by ApoAI amyloidosis. Our study provides the first molecular evidence that a significant fraction of ApoAI amyloidosis cases with no family history result from spontaneous neomutations rather than variable disease penetrance. Finally, successful hepatorenal transplantation resulted in a life- and vision-saving measure for a 32-year-old man with a hitherto unreported severe ApoAI amyloidosis caused by the very rare Glu34Lys variant. Our findings reveal new modes of occurrence and expand the clinical spectrum of ApoAI amyloidosis. The awareness of glomerular and ocular manifestations in ApoAI amyloidosis should enable earlier diagnosis and avoid misdiagnosis with other forms of renal amyloidosis. Thus, documented apoA-I amyloid deposition in the retina offers new biological information about this disease and may change organ transplantation practice to reduce retinal damage in patients with ApoAI amyloidosis.

Renal toxicities associated with pembrolizumab.

OBJECTIVE: Expanded clinical experience with patients treated by pembrolizumab has accumulated. However, renal toxicities associated with this anti-programmed cell death 1 agent are poorly described because kidney histology is rarely sought. As a nephrology referral centre, we aimed to describe the clinic-biological and histopathological characteristics of pembrolizumab-related nephropathy and its response to treatment. METHODS: We conducted a monocentric large case series study, including all pembrolizumab-treated cancer patients presenting a renal toxicity addressed to our centre from 2015 to 2017. RESULTS: A total of 12 patients (7 men) out of 676 pembrolizumab-treated patients (incidence 1.77%) were included (median age 69.75 years). Patients were referred for acute kidney injury (n = 10) and/or proteinuria (n = 2). A kidney biopsy was performed in all patients, with a median duration of use of 9 months (range 1-24 months) after the beginning of treatment. Biopsy showed that four patients had acute interstitial nephritis (AIN), whereas five had acute tubular injury (ATI) alone, one had minimal change disease (MCD) and ATI, and one had MCD alone. Pembrolizumab withdrawal coupled with corticosteroid therapy was the most effective treatment for kidney function recovery. Drug reintroduction resulted in a more severe recurrence of AIN in one patient who required maintenance of pembrolizumab. Two patients died of cancer progression with one of them developing severe renal failure requiring dialysis. CONCLUSION: In our series, ATI, AIN and MCD are the most frequent forms of kidney involvement under pembrolizumab therapy. Kidney dysfunction is usually isolated but can be severe. Use of corticosteroids in case of AIN improves the glomerular filtration rate.

Decoding cold ischaemia time impact on kidney graft: the kinetics of the unfolded protein response pathways.

The relationship between cold ischaemia time (CIT) and adverse outcome is now acknowledged. However, the underlying mechanisms remain to be defined, which slows the development of adapted therapeutics and diagnostics. We explored the impact of CIT in both preclinical and in vitro models of preservation. We determined that the endoplasmic reticulum (ER) and its stress response (unfolded protein response, UPR) were regulated in close association with CIT; the eIF2α-ATF4 pathway was inhibited early (1-8 h) at the detriment of cell survival, while the ATF6 pathway was activated late (12-24 h) and associated with cell death. The IRE1α-XBP1 branch was activated at reperfusion only if CIT extended beyond 8 h, and had a dual role on cell fate - deleterious through IRE1's RNase activity and beneficial through IRE1α other roles. Finally, the pro-apoptotic factor CHOP was a common target of both ATF6 and IRE1α pathways and was associated with elongated CIT and increased cell death. Microarray analysis of human transplanted kidney confirmed that UPR markers were regulated by CIT and that CHOP was associated with adverse outcome. We show that UPR could be a critical pathway explaining the relationship between CIT and graft outcome, highlighting the potential for UPR-based therapeutics and diagnostics to improve transplantation.

Cutibacterium acnes protects Candida albicans from the effect of micafungin in biofilms.

The aim of this study was to investigate the ability of Candida albicans and Cutibacterium acnes to grow together as a polymicrobial biofilm in vitro and to examine the influence of C. acnes on C. albicans susceptibility to micafungin. Mature 72-h-old single-species biofilms of C. albicans and polymicrobial biofilms involving both C. albicans and C. acnes were formed in brain-heart infusion and were observed by scanning electronic microscopy. Moreover, 24-h-old single-species and polymicrobial biofilms were treated for 24 h with micafungin (concentrations ranging from 0.75 mg/L to 12 mg/L) and the antibiofilm activity of micafungin was evaluated on fungal cells by flow cytometry following addition of propidium iodide. The results showed that C. albicans and C. acnes formed a polymicrobial biofilm in the tested conditions and that bacterial presence did not modify fungal viability. Micafungin induced a fungal mortality rate ranging from 70-95% in C. albicans single-species biofilms and from 35-40% in C. acnes-C. albicans polymicrobial biofilms. Mortality induced by micafungin was significantly reduced (P < 0.05 for micafungin at 6 mg/L and P < 0.001 for other micafungin concentrations) in polymicrobial conditions compared with single-species biofilms. In conclusion, this study showed that C. albicans and C. acnes are able to form polymicrobial biofilms together in a synergistic way and that this organisation increases yeast resistance to micafungin.

DNAJB9 Is a Specific Immunohistochemical Marker for Fibrillary Glomerulonephritis.

INTRODUCTION: Fibrillary glomerulonephritis (FGN) is a rare disease with unknown pathogenesis and a poor prognosis. Until now, the diagnosis of this disease has required demonstration of glomerular deposition of randomly oriented fibrils by electron microscopy that are Congo red negative and stain with antisera to Igs. We recently discovered a novel proteomic tissue biomarker for FGN, namely, DNAJB9. METHODS: In this work, we developed DNAJB9 immunohistochemistry and tested its sensitivity and specificity for the diagnosis of FGN. This testing was performed on renal biopsy samples from patients with FGN (n = 84), amyloidosis (n = 21), a wide variety of non-FGN glomerular diseases (n = 98), and healthy subjects (n = 11). We also performed immunoelectron microscopy to determine whether DNAJB9 is localized to FGN fibrils. RESULTS: Strong, homogeneous, smudgy DNAJB9 staining of glomerular deposits was seen in all but 2 cases of FGN. The 2 cases that did not stain for DNAJB9 were unique, as they had glomerular staining for IgG only (without κ or λ) on immunofluorescence. DNAJB9 staining was not observed in cases of amyloidosis, in healthy subjects, or in non-FGN glomerular diseases (with the exception of very focal staining in 1 case of smoking-related glomerulopathy), indicating 98% sensitivity and > 99% specificity. Immunoelectron microscopy showed localization of DNAJB9 to FGN fibrils but not to amyloid fibrils or immunotactoid glomerulopathy microtubules. CONCLUSION: DNAJB9 immunohistochemistry is sensitive and specific for FGN. Incorporation of this novel immunohistochemical biomarker into clinical practice will now allow more rapid and accurate diagnosis of this disease.

Ultrastructural study of liver and lead tissue concentrations in young mallard ducks (Anas platyrhynchos) after ingestion of single lead shot.

Lead (Pb) represents a serious threat to wildlife and ecosystems. The aim of this study was to examine the subcellular effects of dietary Pb pellet ingestion on mallard (Anas platyrhynchos) livers. After ingestion of a single Pb shot (LS4 size class: 0.177 ± 0.03 g) in 41 mallard ducks (22 males and 19 females) versus 10 controls (5 males and 5 females), all 7-week old, a morphologic study was conducted by TEM (transmission electron microscopy) of liver at the subcellular level. The results in treated mallards showed at a magnification of 2500 X that hepatic parenchyma was altered as evidenced by intralysosomal electron-dense deposits, which are compatible with Pb deposits. Further, at a higher magnification (15,000 X) in both genders, deterioration of mitochondria was observed in which the crests and, to a lesser extent, outer membrane were lysed. While the rough endoplasmic reticulum was fragmented, intracytoplasmic electron-dense material compatible with Pb deposits was maximally visible, thereby underscoring the deeply destructive effect of this metal on the subcellular architecture of the liver. In addition, applying an optimized and validated method in a clean room using electrothermal atomic absorption spectrophotometer (ETAAS) with Zeeman background correction, the objective was to improve and refine certain indispensable measurements pertaining to Pb impregnation in tissues other than liver such as kidneys, bones, and feathers of mallards. Data demonstrated show that compared with controls, Pb accumulation increases significantly, not only in the liver (3-fold), but also in the bones and the feathers (14-fold). No significant difference was noted between males and females. Bearing in mind the marked subcellular toxicity attributed to Pb, this study reinforces present-day arguments advocating limitation of game consumption.

An observational study of phagocytes and Klebsiella pneumoniae relationships: different behaviors.

Klebsiella pneumoniae is a bacterium that can be in relation with free living amoebae like Acanthamoeba castellanii in natural environments such as soil and water. This pathogen, which is responsible for community-acquired pneumonia and for nosocomial infections, also has interactions with host defense mechanisms like macrophages. As it has been shown that A. castellanii shares some traits with macrophages, in particular the ability to phagocyte bacteria, we have studied the uptake and the fate of the bacteria after contact with the two phagocytic cells. In our conditions, K. pneumoniae growth was increased in coculture in presence of A. castellanii or Thp-1 macrophagic cells and bacterial development was also increased by A. castellanii supernatant. In addition, we showed that the presence of the bacteria had a negative effect on the macrophages whereas it does not affect amoeba viability. Using gentamicin, which kills bacteria outside cells, we showed that only macrophages were able to internalize K. pneumoniae. This result was confirmed by electron microscopy. We have consequently reported some differences in bacterial uptake and internalization between a free living amoeba and macrophagic cells, highlighting the fact that results obtained with this amoebal model should not be extrapolated to the relationships between K. pneumoniae and macrophages.

Proliferative lupus nephritis in the absence of overt systemic lupus erythematosus: A historical study of 12 adult patients.

Severe lupus nephritis in the absence of systemic lupus erythematosus (SLE) is a rare condition with an unclear clinical presentation and outcome.We conducted a historical observational study of 12 adult (age >18 years) patients with biopsy-proven severe lupus nephritis or lupus-like nephritis without SLE immunological markers at diagnosis or during follow-up. Excluded were patients with chronic infections with HIV or hepatitis B or C; patients with a bacterial infectious disease; and patients with pure membranous nephropathy. Electron microscopy was retrospectively performed when the material was available. End points were the proportion of patients with a complete response (urine protein to creatinine ratio <0.5 g/day and a normal or near-normal eGFR), partial response (≥50% reduction in proteinuria to subnephrotic levels and a normal or near-normal eGFR), or nonresponse at 12 months or later after the initiation of the treatment.The study included 12 patients (66% female) with a median age of 36.5 years. At diagnosis, median creatinine and proteinuria levels were 1.21 mg/dL (range 0.5-11.6) and 7.5 g/day (1.4-26.7), respectively. Six patients had nephrotic syndrome and acute kidney injury. Renal biopsy examinations revealed class III or class IV A/C lupus nephritis in all cases. Electron microscopy was performed on samples from 5 patients. The results showed mesangial and subendothelial dense deposits consistent with LN in 4 cases, and a retrospective diagnosis of pseudo-amyloid fibrillary glomerulonephritis was made in 1 patient.Patients received immunosuppressive therapy consisting of induction therapy followed by maintenance therapy, similar to treatment for severe lupus nephritis. Remission was recorded in 10 patients at 12 months after the initiation of treatment. One patient reached end-stage renal disease. After a median follow-up of 24 months, 2 patients relapsed.Lupus nephritis in the absence of overt SLE is a nosological entity requiring careful etiological investigation, including systematic electron microscopy examination of renal biopsies to rule out fibrillary glomerulonephritis. In this series, most patients presented with severe glomerulonephritis, which was highly similar to lupus nephritis at presentation and in terms of response to immunosuppressive therapy.

In vitro culture and phenotypic and molecular characterization of gastric stem cells from human stomach.

BACKGROUND: Human gastric mucosa shows continuous self-renewal via differentiation from stem cells that remain poorly characterized. METHODS: We describe an original protocol for culture of gastric stem/progenitor cells from adult human stomach. The molecular characteristics of cells were studied using TaqMan low-density array and qRT-PCR analyses using the well-characterized H1 and H9 embryonic stem cells as reference. Epithelial progenitor cells were challenged with H. pylori to characterize their inflammatory response. RESULTS: Resident gastric stem cells expressed specific molecular markers of embryonic stem cells (SOX2, NANOG, and OCT4), as well as others specific to adult stem cells, particularly LGR5 and CD44. We show that gastric stem cells spontaneously differentiate into epithelial progenitor cells that can be challenged with H. pylori. The epithelial progenitor response to H. pylori showed a cag pathogenicity island-dependent induction of matrix metalloproteinases 1 and 3, chemokine (CXCL1, CXCL5, CXCL8, CCL20) and interleukine 33 expression. CONCLUSION: This study opens new outlooks for investigation of gastric stem cell biology and pathobiology as well as host-H. pylori interactions.

Unravelling the immunopathological mechanisms of heavy chain deposition disease with implications for clinical management.

Randall-type heavy chain deposition disease (HCDD) is a rare disorder characterized by tissue deposition of a truncated monoclonal immunoglobulin heavy chain lacking the first constant domain. Pathophysiological mechanisms are unclear and management remains to be defined. Here we retrospectively studied 15 patients with biopsy-proven HCDD of whom 14 presented with stage 3 or higher chronic kidney disease, with nephrotic syndrome in 9. Renal lesions were characterized by nodular glomerulosclerosis, with linear peritubular and glomerular deposits of γ-heavy chain in 12 patients or α-heavy chain in 3 patients, without concurrent light chain staining. Only 2 patients had symptomatic myeloma. By serum protein electrophoresis/immunofixation, 13 patients had detectable monoclonal gammopathy. However, none of these techniques allowed detection of the nephrotoxic truncated heavy chain, which was achieved by immunoblot and/or bone marrow heavy chain sequencing in 14 of 15 patients. Serum-free kappa to lambda light chain ratio was abnormal in 11 of 11 patients so examined. Immunofluorescence studies of bone marrow plasma cells showed coexpression of the pathogenic heavy chain with light chain matching the abnormal serum-free light chain in all 3 tested patients. Heavy chain sequencing showed first constant domain deletion in 11 of 11 patients, with high isoelectric point values of the variable domain in 10 of 11 patients. All patients received chemotherapy, including bortezomib in 10 cases. Renal parameters improved in 11 patients who achieved a hematological response, as assessed by normalization of the free light chain ratio in 8 cases. Tissue deposition in HCDD relates to physicochemical peculiarities of both variable and constant heavy chain domains. Early diagnosis and treatment with bortezomib-based combinations appear important to preserve renal prognosis. Thus, monitoring of serum-free light chain is an indirect but useful method to evaluate the hematological response.

Blocking TGF-ß Signaling Pathway Preserves Mitochondrial Proteostasis and Reduces Early Activation of PDGFRß+ Pericytes in Aristolochic Acid Induced Acute Kidney Injury in Wistar Male Rats.

BACKGROUND: The platelet-derived growth factor receptor ß (PDGFRß)+ perivascular cell activation becomes increasingly recognized as a main source of scar-associated kidney myofibroblasts and recently emerged as a new cellular therapeutic target. AIMS: In this regard, we first confirmed the presence of PDGFRß+ perivascular cells in a human case of end-stage aristolochic acid nephropathy (AAN) and thereafter we focused on the early fibrosis events of transforming growth factor ß (TGFß) inhibition in a rat model of AAN. MATERIALS AND METHODS: Neutralizing anti-TGFß antibody (1D11) and its control isotype (13C4) were administered (5 mg/kg, i.p.) at Days -1, 0, 2 and 4; AA (15 mg/kg, sc) was injected daily. RESULTS: At Day 5, 1D11 significantly suppressed p-Smad2/3 signaling pathway improving renal function impairment, reduced the score of acute tubular necrosis, peritubular capillaritis, interstitial inflammation and neoangiogenesis. 1D11 markedly decreased interstitial edema, disruption of tubular basement membrane loss of brush border, cytoplasmic edema and organelle ultrastructure alterations (mitochondrial disruption and endoplasmic reticulum edema) in proximal tubular epithelial cells. Moreover, 1D11 significantly inhibited p-PERK activation and attenuated dysregulation of unfolded protein response (UPR) pathways, endoplasmic reticulum and mitochondrial proteostasis in vivo and in vitro. CONCLUSIONS: The early inhibition of p-Smad2/3 signaling pathway improved acute renal function impairment, partially prevented epithelial-endothelial axis activation by maintaining PTEC proteostasis and reduced early PDGFRß+ pericytes-derived myofibroblasts accumulation.

Prognostic value of kidney biopsy in myeloma cast nephropathy: a retrospective study of 70 patients.

BACKGROUND: Light chain myeloma cast nephropathy (MCN) is the major cause of renal failure in multiple myeloma and strongly impacts patient survival. The role of kidney biopsy in the management of MCN is unclear. METHODS: Renal pathological findings were retrospectively studied in 70 patients with multiple myeloma and MCN. Patients were categorized according to the achievement or not of renal response, as defined by estimated glomerular filtration rate (eGFR) ≥ 30 mL/min/1.73 m(2) and/or dialysis independence at 3 months. RESULTS: Thirty-two patients (46%) achieved a renal response. In the whole study population, the following parameters differed significantly between patients with and without renal response, respectively: baseline median eGFR (13.3 versus 9.3 mL/min/1.73 m(2), P = 0.017), Acute Kidney Injury Network Stage 3 (68.8 versus 92.1%, P = 0.019), haematological response rate (94 versus 34%, P < 0.0001), median percentage of free light chain (FLC) reduction at Day 21 (92 versus 24%, P = 0.006) and median number of casts/10 fields (14 versus 25, P = 0.005). The extent of interstitial fibrosis and tubular atrophy was similar. In multivariate analysis, only FLC reduction at Day 21 was significantly associated with renal response. However, when considering only the subgroup of haematological responders, both median number of casts [odds ratio (OR) = 0.93, 95% confidence interval (95% CI): 0.88-0.98, P = 0.01] and extent of tubular atrophy (OR = 0.03, 95% CI: 0.00-0.52, P = 0.02) were independent predictors of renal response. CONCLUSIONS: In MCN, the presence of numerous casts and diffuse tubular atrophy is associated with poor renal prognosis. These data suggest that additional strategies to reduce FLC burden should be considered in patients with extensive cast formation.

Activity of Six Essential Oils Extracted from Tunisian Plants against Legionella pneumophila.

The aim of this study was to investigate the composition of six essential oils extracted from Tunisian plants, i.e., Artemisia herba-alba Asso, Citrus sinensis (L.) Osbeck, Juniperus phoenicea L., Rosmarinus officinalis L., Ruta graveolens L., and Thymus vulgaris L., and to evaluate their activity against Legionella pneumophila (microdilution assays). Eight Legionella pneumophila strains were studied, including the two well-known serogroup 1 Lens and Paris strains as controls and six environmental strains isolated from Tunisian spas belonging to serogroups 1, 4, 5, 6, and 8. The essential oils were generally active against L. pneumophila. The activities of the A. herba-alba, C. sinensis, and R. officinalis essential oils were strain-dependent, whereas those of the J. phoenicea and T. vulgaris oils, showing the highest anti-Legionella activities, with minimum inhibitory concentrations (MICs) lower than 0.03 and lower than or equal to 0.07 mg/ml, respectively, were independent of the strains' serogroup. Moreover, the microorganisms treated with T. vulgaris essential oil were shorter, swollen, and less electron-dense compared to the untreated controls. Isoborneol (20.91%), (1S)-α-pinene (18.30%) ß-phellandrene (8.08%), α-campholenal (7.91%), and α-phellandrene (7.58%) were the major components isolated from the J. phoenicea oil, while carvacrol (88.50%) was the main compound of the T. vulgaris oil, followed by p-cymene (7.86%). This study highlighted the potential interest of some essential oils extracted from Tunisian plants as biocides to prevent the Legionella risk.

A mouse model recapitulating human monoclonal heavy chain deposition disease evidences the relevance of proteasome inhibitor therapy.

Randall-type heavy chain deposition disease (HCDD) is a rare disorder characterized by glomerular and peritubular amorphous deposits of a truncated monoclonal immunoglobulin heavy chain (HC) bearing a deletion of the first constant domain (CH1). We created a transgenic mouse model of HCDD using targeted insertion in the immunoglobulin κ locus of a human HC extracted from a HCDD patient. Our strategy allows the efficient expression of the human HC in mouse B and plasma cells, and conditional deletion of the CH1 domain reproduces the major event underlying HCDD. We show that the deletion of the CH1 domain dramatically reduced serum HC levels. Strikingly, even with very low serum level of truncated monoclonal HC, histologic studies revealed typical Randall-type renal lesions that were absent in mice expressing the complete human HC. Bortezomib-based treatment resulted in a strong decrease of renal deposits. We further demonstrated that this efficient response to proteasome inhibitors mostly relies on the presence of the isolated truncated HC that sensitizes plasma cells to bortezomib through an elevated unfolded protein response (UPR). This new transgenic model of HCDD efficiently recapitulates the pathophysiologic features of the disease and demonstrates that the renal damage in HCDD relies on the production of an isolated truncated HC, which, in the absence of a LC partner, displays a high propensity to aggregate even at very low concentration. It also brings new insights into the efficacy of proteasome inhibitor-based therapy in this pathology.

Kidney diseases associated with monoclonal immunoglobulin M-secreting B-cell lymphoproliferative disorders: a case series of 35 patients.

BACKGROUND: Kidney diseases associated with immunoglobulin M (IgM) monoclonal gammopathy are poorly described, with few data for patient outcomes and renal response. STUDY DESIGN: Case series. SETTING & PARTICIPANTS: 35 patients from 8 French departments of nephrology were retrospectively studied. Inclusion criteria were: (1) detectable serum monoclonal IgM, (2) estimated glomerular filtration rate (eGFR) < 60mL/min/1.73m(2) and/or proteinuria with protein excretion > 0.5g/d and/or microscopic hematuria, and (3) kidney biopsy showing monoclonal immunoglobulin deposits and/or lymphomatous B-cell renal infiltration. All patients received chemotherapy, including rituximab-based regimens in 8 cases. PREDICTORS: Patients were classified into 3 groups according to renal pathology: glomerular AL amyloidosis (group 1; n=11), nonamyloid glomerulopathies (group 2; n=15, including 9 patients with membranoproliferative glomerulonephritis), and tubulointerstitial nephropathies (group 3; n=9, including cast nephropathy in 5, light-chain Fanconi syndrome in 3, and isolated tumor infiltration in 1). OUTCOMES: Posttreatment hematologic response (≥50% reduction in serum monoclonal IgM and/or free light chain level) and renal response (≥50% reduction in 24-hour proteinuria or eGFR≥30mL/min/1.73m(2) in patients with glomerular and tubulointerstitial disorders, respectively). RESULTS: Nephrotic syndrome was observed in 11 and 6 patients in groups 1 and 2, respectively. Patients in group 3 presented with acute kidney injury (n=7) and/or proximal tubular dysfunction (n=3). Waldenström macroglobulinemia was present in 26 patients (8, 12, and 6 in groups 1, 2, and 3, respectively). Significant lymphomatous interstitial infiltration was observed in 18 patients (4, 9, and 5 patients, respectively). Only 9 of 29 evaluable patients had systemic signs of symptomatic hematologic disease (2, 5, and 2, respectively). In groups 1, 2, and 3, respectively, hematologic response was achieved after first-line treatment in 3 of 9, 9 of 10, and 5 of 6 evaluable patients, while renal response occurred in 5 of 10, 9 of 15, and 5 of 8 evaluable patients. LIMITATIONS: Retrospective study; insufficient population to establish the impact of chemotherapy. CONCLUSIONS: IgM monoclonal gammopathy is associated with a wide spectrum of renal manifestations, with an under-recognized frequency of tubulointerstitial disorders.

Morphological Study of the Encystment and Excystment of Vermamoeba vermiformis Revealed Original Traits.

Free-living amoebae are ubiquitous protozoa commonly found in water. Among them, Acanthamoeba and Vermamoeba (formerly Hartmannella) are the most represented genera. In case of stress, such as nutrient deprivation or osmotic stress, these amoebae initiate a differentiation process, named encystment. It leads to the cyst form, which is a resistant form enabling amoebae to survive in harsh conditions and resist disinfection treatments. Encystment has been thoroughly described in Acanthamoeba but poorly in Vermamoeba. Our study was aimed to follow the encystment/excystment processes by microscopic observations. We show that encystment is quite rapid, as mature cysts were obtained in 9 h, and that cyst wall is composed of two layers. A video shows that a locomotive form is likely involved in clustering cysts together during encystment. As for Acanthamoeba, autophagy is likely active during this process. Specific vesicles, possibly involved in ribophagy, were observed within the cytoplasm. Remarkably, mitochondria rearranged around the nucleus within the cyst, suggesting high needs in energy. Unlike Acanthamoeba and Naegleria, no ostioles were observed in the cyst wall suggesting that excystment is original. During excystment, large vesicles, likely filled with hydrolases, were found in close proximity to cyst wall and digest it. Trophozoite moves inside its cyst wall before exiting during excystment. In conclusion, Vermamoeba encystment/excystment displays original trends as compare to Acanthamoeba.

Stenotrophomonas maltophilia and Vermamoeba vermiformis relationships: bacterial multiplication and protection in amoebal-derived structures.

Stenotrophomonas maltophilia, a bacteria involved in healthcare-associated infections, can be found in hospital water systems. Other microorganisms, such as Free Living amoebae (FLA), are also at times recovered in the same environment. Amongst these protozoa, many authors have reported the presence of Vermamoeba vermiformis. We show here that this amoeba enhances S. maltophilia growth and harbors the bacteria in amoebal-derived structures after 28 days in harsh conditions. These results highlight the fact that particular attention should be paid to the presence of FLA in hospital water systems, because of their potential implication in survival and growth of pathogenic bacterial species.

Impairment of autophagy in the central nervous system during lipopolysaccharide-induced inflammatory stress in mice.

BACKGROUND: Current evidence suggests a central role for autophagy in many neurodegenerative diseases including Alzheimer's disease, Huntington's disease, Parkinson's disease and amyotrophic lateral sclerosis. Furthermore, it is well admitted that inflammation contributes to the progression of these diseases. Interestingly, crosstalks between autophagy and inflammation have been reported in vitro and at the peripheral level such as in Crohn's disease. However, the impact of systemic inflammation on autophagic components in the brain remains to be documented. Therefore, this study monitored autophagy markers after acute and chronic lipopolysaccharide (LPS)-induced inflammatory stress in mice. RESULTS: We showed that acute inflammation, 24 h post-intraperitoneal 10 mg/kg LPS, substantially increased cytokine production (Interleukin(IL)-1ß, Tumor necrosis factor (TNF)-α and IL-6), decreased the levels of autophagy markers (Beclin-1, p62 and LC3 II) and reduced p70S6K activation in cortex and hippocampus. In hippocampus, IL-1ß levels and LC3 II expression were positively and highly correlated and a negative correlation was noted between TNF-α levels and p70S6K activation. Chronic inflammation by injection of 0.5 mg/kg LPS every three days during three months led to a moderate IL-1ß production and decreased TNF-α levels. Interestingly, Beclin-1 and LC3 II levels decreased while those of p62 increased. Cortical IL-1ß levels positively correlated with Beclin-1 and LC3 II and on the contrary inversely correlated with p62. CONCLUSION: The present study is the first showing links between IL-1ß-mediated inflammation and autophagy in the brain. It could open to new therapeutic strategies in brain diseases where regulation impairment of inflammation and autophagy progress with the severity of diseases.