M-30 and 4HNE are sequestered in different aggresomes in the same hepatocytes

M-30 and 4HNE adducts are two markers of active liver disease. M-30 is a serologic marker and 4HNE adducts are histologic markers. M-30 is a marker for apoptosis because it is a fragment of cytokeratin-18 left over from proteolysis by caspase 3. 4HNE is a marker of oxidative stress because it results from lipid peroxidation. Both markers are commonly found in nonalcoholic steatohepatitis and in alcoholic hepatitis. Liver biopsies from patients with steatohepatitis, 11 alcoholic and 11 non-alcoholics were stained for 4HNE and M-30. Almost all of the biopsies in both groups showed 4HNE- and M-30-positive aggresomes in hepatocytes. Mallory Denk bodies (MDB) stained variably positive for M-30, whereas 4HNE was present in aggresomes independent of MDBs. However, they were sometimes located in hepatocytes which also contained MDBs as shown by confocal microscopy of double stained biopsies. The results indicate that the formation of M-30 and 4HNE aggresomes occurs through different pathways of liver cell injury in both types of steatohepatitis.     

Side‐effects of thiamethoxam on the brain andmidgut of the africanized honeybee Apis mellifera (Hymenopptera: Apidae)

The development of agricultural activities coincides with the increased use of pesticides to control pests, which can also be harmful to nontarget insects such as bees. Thus, the goal of this work was assess the toxic effects of thiamethoxam on newly emerged worker bees of Apis mellifera (africanized honeybee—AHB). Initially, we determined that the lethal concentration 50 (LC₅₀) of thiamethoxam was 4.28 ng a.i./μL of diet. To determine the lethal time 50 (LT₅₀), a survival assay was conducted using diets containing sublethal doses of thiamethoxam equal to 1/10 and 1/100 of the LC_50. The group of bees exposed to 1/10 of the LC₅₀ had a 41.2% reduction of lifespan. When AHB samples were analyzed by morphological technique we found the presence of condensed cells in the mushroom bodies and optical lobes in exposed honeybees. Through Xylidine Ponceau technique, we found cells which stained more intensely in groups exposed to thiamethoxam. The digestive and regenerative cells of the midgut from exposed bees also showed morphological and histochemical alterations, like cytoplasm vacuolization, increased apocrine secretion and increased cell elimination. Thus, intoxication with a sublethal doses of thiamethoxam can cause impairment in the brain and midgut of AHB and contribute to the honeybee lifespan reduction. © 2013 Wiley Periodicals, Inc. Environ Toxicol 29: 1122–1133, 2014.     

Type II Grass Carp Reovirus Infects Leukocytes but Not Erythrocytes and Thrombocytes in Grass Carp (Ctenopharyngodon idella)

Grass carp reovirus (GCRV) causes serious losses to the grass carp industry. At present, infectious tissues of GCRV have been studied, but target cells remain unclear. In this study, peripheral blood cells were isolated, cultured, and infected with GCRV. Using quantitative real-time polymerase chain reaction (qRT-PCR), Western Blot, indirect immunofluorescence, flow cytometry, and transmission electron microscopy observation, a model of GCRV infected blood cells in vitro was established. The experimental results showed GCRV could be detectable in leukocytes only, while erythrocytes and thrombocytes could not. The virus particles in leukocytes are wrapped by empty membrane vesicles that resemble phagocytic vesicles. The empty membrane vesicles of leukocytes are different from virus inclusion bodies in C. idella kidney (CIK) cells. Meanwhile, the expression levels of IFN1, IL-1β, Mx2, TNFα were significantly up-regulated in leukocytes, indicating that GCRV could cause the production of the related immune responses. Therefore, GCRV can infect leukocytes in vitro, but not infect erythrocytes and thrombocytes. Leukocytes are target cells in blood cells of GCRV infections. This study lays a theoretical foundation for the study of the GCRV infection mechanism and anti-GCRV immunity.     

Addition of MHPG to Alzheimer's disease biomarkers improves differentiation of dementia with Lewy bodies from Alzheimer's disease but not other dementias

BackgroundOverlapping clinical features make it difficult to distinguish dementia with Lewy bodies (DLB) from Alzheimer's disease (AD) and other dementia types. In this study we aimed to determine whether the combination of cerebrospinal fluid (CSF) biomarkers, amyloid-β₄₂ (Aβ₄₂), total tau protein (t-tau), and phosphorylated tau protein (p-tau), in combination with 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG), could be useful in discriminating DLB from vascular dementia (VaD) and frontotemporal dementia (FTD), as we previously demonstrated for differentiation of DLB from AD.MethodsWe retrospectively analyzed concentrations of MHPG, Aβ₄₂, t-tau, and p-tau in CSF in patients with DLB, AD, VaD, and FTD. Using previously developed multivariate logistic regression models we assessed the diagnostic value of these CSF parameters.ResultsThe currently used combination of Aβ₄₂, t-tau, and p-tau yielded a sensitivity of 61.9% and a specificity of 91.7% for the discrimination between DLB and AD, but could not discriminate between DLB and VaD or FTD. The addition of MHPG to Aβ₄₂, t-tau, and p-tau improves the discrimination of DLB from AD, yielding a sensitivity of 65.1% and specificity of 100%, but could not distinguish DLB from other forms of dementia.ConclusionsOur results confirm in a separate patient cohort that addition of MHPG to Aβ₄₂, t-tau, and p-tau improves the discrimination of DLB from AD but not the differentiation of DLB from VaD or FTD.     

Beta cyclodextrins bind,stabilize, and remove lipofuscin bisretinoids from retinal pigment epithelium

Accumulation of lipofuscin bisretinoids (LBs) in the retinal pigment epithelium (RPE) is the alleged cause of retinal degeneration in genetic blinding diseases (e.g., Stargardt) and a possible etiological agent for age-related macular degeneration. Currently, there are no approved treatments for these diseases; hence, agents that efficiently remove LBs from RPE would be valuable therapeutic candidates. Here, we show that beta cyclodextrins (β-CDs) bind LBs and protect them against oxidation. Computer modeling and biochemical data are consistent with the encapsulation of the retinoid arms of LBs within the hydrophobic cavity of β-CD. Importantly, β-CD treatment reduced by 73% and 48% the LB content of RPE cell cultures and of eyecups obtained from Abca4-Rdh8 double knock-out (DKO) mice, respectively. Furthermore, intravitreal administration of β-CDs reduced significantly the content of bisretinoids in the RPE of DKO animals. Thus, our results demonstrate the effectiveness of β-CDs to complex and remove LB deposits from RPE cells and provide crucial data to develop novel prophylactic approaches for retinal disorders elicited by LBs.The retinal pigment epithelium (RPE), strategically situated between the neural retina and the choroid blood vessels, is essential for photoreceptor (PR) function. It recycles vitamin A, which is required for the visual cycle and clears debris generated by the circadian shedding of PR outer segments (1, 2). Each RPE cell phagocytoses and digests the material produced by 30–50 overlying PRs, which shed 10% of their mass daily. The intense and continual phagocytic activity of RPE cells results in the progressive accumulation of indigestible products or “lipofuscin” in their lysosomal compartment (3, 4). Unlike lipofuscins found in other body tissues, which are composed mainly of protein, RPE lipofuscin consists predominantly of lipid-bisretinoids and only 2% protein (5). Lipofuscin bisretinoids (LBs) are vitamin A-derived side products of the visual cycle. Light converts 11-cis-retinal (11CR), the visual pigment chromophore, into all-trans-retinal (ATR), which is immediately flipped by the ATP-binding cassette transporter 4 (Abca4) transporter from the lumen of the outer segment discs to the cytoplasm, where it is reduced to inert all-trans-retinol by retinol dehydrogenase 8 (Rdh8), in mice (6, 7). Small fractions of 11CR and ATR are converted into N-retinylidine-N-ethanolamine (A2E) and other less abundant bisretinoids, which once accumulated in the lysosomes of RPE cells are refractory to all known lysosomal hydrolases (8, 9). The concept that LB accumulation causes retinal degeneration is supported by in vitro and in vivo data that show that excessive LBs are toxic for cultured RPE cells (10, 11), that photoreceptors overlying A2E-laden RPE are more prone to degeneration (12) and that excessive accumulation of LBs in Stargardt’s disease precedes macular degeneration (13). Mice carrying null mutations in Abca4 and Rdh8 develop blindness, basal laminar deposits, and focal accumulations of extracellular debris between the RPE and the Bruch membrane (drusen) (6).Here we report that a family of modified cyclic oligosaccharides, beta cyclodextrins (β-CDs), formed by seven d-glucose units, can encapsulate the hydrophobic arms of A2E within their nonpolar cavity, protect A2E from oxidation, and remove A2E from RPE cells. Our data demonstrate a direct correlation between the ability of β-CDs to perform these protective functions and their affinity for A2E.     

The PML domain of PML–RARα blocks senescence to promote leukemia

In most acute promyelocytic leukemia (APL) cases, translocons produce a promyelocytic leukemia protein–retinoic acid receptor α (PML–RARα) fusion gene. Although expression of the human PML fusion in mice promotes leukemia, its efficiency is rather low. Unexpectedly, we find that simply replacing the human PML fusion with its mouse counterpart results in a murine PML–RARα (mPR) hybrid protein that is transformed into a significantly more leukemogenic oncoprotein. Using this more potent isoform, we show that mPR promotes immortalization by preventing cellular senescence, impeding up-regulation of both the p21 and p19^ARF cell-cycle regulators. This induction coincides with a loss of the cancer-associated ATRX/Daxx–histone H3.3 predisposition complex and suggests inhibition of senescence as a targetable mechanism in APL therapy.Acute promyelocytic leukemia (APL) is characterized by chromosomal translocations involving retinoic acid receptor alpha (RARα) with a limited number of translocation partners. A common feature of APL-promoting fusion proteins is their ability to self-associate. Indeed, previous studies have shown that fusion of RARα with self-associating domains is sufficient to render RARα leukemogenic (1). In APL patients, the predominant leukemogenic protein found in 95–99% of cases is the result of the fusion of promyelocytic leukemia protein (PML) with RARα (human PML–RARα; hPR) (2, 3). RARα and PML are regulatory proteins implicated in multiple aspects of differentiation and development (4) and apoptosis and cellular senescence (5, 6), respectively. Despite speculation, the relevance of senescence in APL is not fully understood (7, 8).Current mouse models recapitulate many key features of the human disease, including a malignant promyelocytic phenotype and sensitivity to all-trans retinoic acid (ATRA), but suffer from incomplete penetrance and long latency until disease presentation (1, 9, 10). We reasoned that the relatively low leukemogenic activity of hPR in mice might be due to modest sequence identity between human and mouse PML (PML: 63% identity; RARα: 98% identity). Consistent with this notion, we have designed an “experimental oncoprotein” corresponding to the fusion of mouse PML with RARα (mPR), which produced myelocytic leukemia similar to hPR-induced murine APL (10) but with higher penetrance and shorter latency periods. Notably, expression of mPR disrupted PML nuclear bodies (PML-NBs), phenocopying hPR-induced APL (11, 12). We show here that senescence-related up-regulation of p21 and p19 is completely lost in primary murine bone marrow cells upon expression of mPR. Furthermore, we find that the assembly of the death domain associated protein (Daxx)–alpha thalassemia/mental retardation syndrome X-linked (ATRX) complex at PML-NBs is disrupted by mPR expression, implicating this PML–ATRX–Daxx (PAX) complex in cellular senescence and tumor suppressor activity for PML (13). This study provides experimental evidence for the relevance of PML-NB disruption in APL genesis.     

Dementia with Lewy Bodies with Pure Agraphia for Kanji (Japanese Morphograms)

A 70-year-old dextral woman was admitted to a hospital with agraphia for kanji (Japanese morphograms). She had a history of severe constipation, nightmares, and visual hallucinations. Neurological examinations revealed no obvious Parkinson''s disease symptoms. She showed poor skills in writing the kanji for looking at picture objects, [e.g., writing the Japanese word “inu” (which means dog) when she saw a drawing of a dog] or dictated words. A reduced striatal uptake of [¹²³I]-2β-carbomethoxy-3β-(4-iodophenyl)-N-(3-fluoropropyl) nortropane (FP-CIT) on single-photon-emission computed tomography and reduced meta-iodobenzylguanidine (MIBG) cardiac uptake on myocardial scintigraphy were detected. The accumulation of amyloid beta in the bilateral cerebral cortices was observed on amyloid-positron emission tomography. We herein report a case of Lewy body dementia with pure agraphia for kanji with underlying Alzheimer''s disease pathology.     

A curious case of foreign body induced jejunal obstruction and perforation

INTRODUCTION

Accidental and unnoticed ingestion of foreign bodies are not very uncommon. Most of such foreign bodies pass through gastrointestinal tract uneventfully and only on rare instances cause obstruction and/or perforation.

PRESENTATION OF CASE

We are reporting a case in which a 45 yr old male presented to accident and emergency department of our institute with complaints of pain abdomen, greenish vomiting, abdominal distension, fever and non passage of flatus and stool following alcoholic binge 15 days back. On presentation tachycardia, hypotension, generalised abdominal tenderness and guarding were present. After initial resuscitation and investigation diagnosis of perforation peritonitis was made and emergency exploratory lapratomy was done and a single perforation with plastic foreign body protruding through it was found in jejunum 5 cm distal to duodeno-jejunal junction.

DISCUSSION

The majority of ingested FBs that reach the stomach pass uneventfully through the gastrointestinal tract. The majority of cases occur in children. Only 1% of patient of patients requires surgical intervention depending upon nature, size and shape of the foreign body.

CONCLUSION

Present case report intends to draw the attention towards possibility of intestinal obstruction and perforation by a single plastic foreign body. High index of suspicion is needed as this foreign body is not even radio-opaque and cannot be picked up in X-ray investigations. Also with increasing use of such plastic materials there are increased chances of such incidents.     

Vesical calculus formation on non-absorbable sutures used for open inguinal hernia repair

INTRODUCTION

Iatrogenic injuries to the urogenital tract are rare, with the bladder being the organ most affected. We describe a case of a vesical calculus that formed on non-absorbable sutures that were used to repair an inguinal hernia.

PRESENTATION OF CASE

A 45-year-old male presented with frank haematuria and dysuria 2 years following an open left inguinal hernia repair. A CT urography showed a vesical calculus adherent to the left anterio-lateral wall of the bladder. Cystoscopy revealed that the calculus formed on non-absorbable sutures. Cystolapaxy was performed followed by cystoscopic excision of the sutures. The patient''s post-operative course was uneventful.

DISCUSSION

Foreign bodies in the urinary bladder always act as a nidus for formation of a calculus. Iatrogenic bladder injuries are common during hernia repair. It is however rare for sutures used to repair an inguinal hernia to involve the urinary bladder wall. The patient most likely had a full bladder at the time of hernia repair or the bladder was part of the contents of the hernia sac.

CONCLUSION

This case illustrates the need to ensure that the bladder is empty prior to pelvic surgery and for surgeons to have a good understanding of inguinal anatomy to avoid injuring the contents of the hernia sac.     

Immunohistochemical localization of spatacsin in α‐synucleinopathies

Spatacsin (SPG11) is a major mutated gene in autosomal recessive spastic paraplegia with thin corpus callosum (ARHSP‐TCC) and is responsible for juvenile Parkinsonism. To elucidate the role of spatacsin in the pathogenesis of α‐synucleinopathies, an immunohistochemical investigation was performed on the brain of patients with Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA) using anti‐spatacsin antibody. In PD, Lewy bodies (LBs) in the brain stem were positive for spatacsin. These LBs showed intense staining in their peripheral portions and occasionally in the central cores. Lewy neurites were also spatacsin‐positive. In DLB, cortical LBs were immunolabeled by spatacsin. In MSA, glial cytoplasmic inclusions (GCI) and a small fraction of neuronal cytoplasmic inclusions (NCI) were positive for spatacsin. The widespread accumulation of spatacsin observed in pathologic α‐synuclein‐containing inclusions suggests that spatacsin may be involved in the pathogenesis of α‐synucleinopathies.