Congo Red Staining Research Articles

Insight Into Factors Influencing the Aggregation Process in Wild-Type and P66R Mutant SOD1: Computational andSpectroscopic Approaches.

Disturbances in metal ion homeostasis associated with amyotrophic lateral sclerosis (ALS) have been described for several years, but the exact mechanism of involvement is not well understood. To elucidate the role of metalation in superoxide dismutase (SOD1) misfolding and aggregation, we comprehensively characterized the structural features (apo/holo forms) of WT-SOD1 and P66R mutant in loop IV. Using computational and experimental methodologies, we assessed the physicochemical properties of these variants and their correlation with protein aggregation at the molecular level. Modifications in apo-SOD1 compared to holo-SOD1 were more pronounced in flexibility, stability, hydrophobicity, and intramolecular interactions, as indicated by molecular dynamics simulations. The enzymatic activities of holo/apo-WT SOD1 were 1.30 and 1.88-fold of the holo/apo P66R mutant, respectively. Under amyloid-inducing conditions, decreased ANS fluorescence intensity in the apo-form relative to the holo-form suggested pre-fibrillar species and amyloid aggregate growth due to occluded hydrophobic pockets. FTIR spectroscopy revealed that apo-WT-SOD1 and apo-P66R exhibited a mixture of parallel and intermolecular β-sheet structures, indicative of aggregation propensity. Aggregate species were identified using TEM, Congo red staining, and ThT/ANS fluorescence spectroscopy. Thermodynamic analyses with GdnHCl demonstrated that metal deficit, mutation, and intramolecular disulfide bond reduction are essential for initiating SOD1 misfolding and aggregation. These disruptions destabilize the dimer-monomer equilibrium, promoting dimer dissociation into monomers and decreasing the thermodynamic stability of SOD1 variants, thus facilitating amyloid/amorphous aggregate formation. Our findings offer novel insights into protein aggregation mechanisms in disease pathology and highlight potential therapeutic strategies against toxic protein aggregation, including SOD1.

A novel dual-staining method for cost-effective visualization and differentiation of microbial biofilms

Microbial biofilms are intricate communities that pose significant challenges in clinical and microbiological settings due to their resistance to antibiotics and immune responses. Advanced microscopy techniques, such as scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), and fluorescence microscopy, are often employed to visualize and differentiate between these biofilms. However, these methods are not feasible in all laboratories because of their high cost and complexity. In contrast, simpler techniques like crystal violet and Congo red staining fail to differentiate bacterial cells from the biofilm matrix. This study introduces a novel dual-staining method using Maneval’s stain for microbial biofilm detection and differentiation. This simple, cost-effective method requires only basic equipment and minimal reagents, making it suitable for routine use across various settings. We applied the dual-staining method to various microbial species, including Staphylococcus aureus, Enterococcus faecalis, Candida albicans, Escherichia coli, and Pseudomonas aeruginosa. When compared with the microtiter plate assay, results showed strong agreement, with the dual-staining method effectively differentiating between bacterial cells and the surrounding biofilm matrix, displaying a distinctive blue polysaccharide layer surrounding the magenta‒red bacterial cells. This technique offers a viable alternative to more expensive and complex biofilm detection methods, with potential applications in clinical diagnostics and biofilm research.

Thymoquinone inhibits Neuroinflammatory mediators and vasoconstriction injury via NF-κB dependent NeuN/GFAP/Ki-67 in hypertensive Dams and F1 male pups on exposure to a mixture of Bisphenol-A analogues

Bisphenol-A (BPA) analogues seem inevitable components of numerous domestic products, but these have been identified as agents of teratogenic disorders. This study, therefore, investigated the effect of thymoquinone (TMQ) on the striatum of hypertensive female rats and their F1 male offsprings, on exposure to a mixture of Bisphenol-B, Bisphenol-F and Bisphenol-S (MBFS). Female rats were divided into normotensive and hypertensive groups; and both were treated with MBFS only, MBFS + TMQ, and TMQ only. Exposure to MBFS and co-treatment with TMQ lasted at least 63 days. Neurobehavioural assessments were conducted using Open Field (OF). A spectrophotometer was used for cholinergic, dopaminergic and adenosinergic enzyme assays; Real-Time PCR for gene expression; and immunohistochemistry for protein quantification; while H&E, cresyl fast violent, and congo red stains were used for histological assessments. From the results, maternal exposure to MBFS mediated striatal dysfunction via p53 and NF-kB upregulation; decreased BCl-2, Ki-67 and NeuN; increased GFAP, nissl bodies and β-amyloid. Dysregulation of cholinergic, dopaminergic and adenosinergic enzymes in addition to decreased nitric oxide levels were also associated with MBFS toxicity. Hypertension was found to exacerbate MBFS toxicity. From OF test; increased anxiety and decreased psychomotor activity were associated with maternal exposure to MBFS. However, co-treatment with thymoquinone prevented striatal dysfunction in hypertensive dams and their F1 male offspring. In conclusion, disruption of the delicate balance between apoptosis and cell proliferation culminating in the reduction of mature neurons is responsible for neurodegeneration and neuropathy associated with MBFS exposure. However, these can be prevented through regular consumption of natural products and supplements rich in thymoquinone.

Digestive Amyloidosis Trends: Clinical, Pathological, and Imaging Characteristics.

Amyloidosis is a group of diseases characterized by the extracellular deposition of abnormally folded, insoluble proteins that lead to organ dysfunction. While it commonly affects the cardiovascular system, gastrointestinal (GI) tract involvement is undetermined. Recent research has focused on understanding the pathophysiology, diagnostic challenges, and therapeutic approaches to GI amyloidosis, particularly in systemic amyloid light-chain (AL) and amyloid A (AA) forms. GI manifestations can include motility disorders, bleeding, and, in severe cases, bowel obstruction. This review highlights the importance of the early recognition of digestive symptoms and associated imagistic findings in GI amyloidosis by analyzing the research that included clinical, pathological, and endoscopic approaches to amyloidosis. A systematic search of the PubMed and Scopus databases identified 19 relevant studies. Our findings showed that amyloid deposits commonly affect the entire GI tract, with AL amyloidosis being the most predominant form. Endoscopic evaluations and biopsy remain key diagnostic tools, with Congo Red staining and mass spectrometry being used to confirm amyloid type. Although progress has been made in diagnosis, the absence of targeted therapies and the indistinct nature of GI symptoms continue to be challenging.

Magnolol acts as a neurorestorative agent in an Aβ1‑42‑induced mouse model of Alzheimer's disease.

Magnolol may have the potential to alleviate the progression of Alzheimer's disease (AD). The present study was conducted to investigate the broader mechanism of action of magnolol in AD pathogenesis. C57BL/6 mice were randomly divided into five groups (n=6 mice/group): i) Control; ii) AD model; iii) 5 mg/kg magnolol + AD model; iv) 10 mg/kg magnolol + AD model; and v) 20 mg/kg magnolol + AD model. A total of 7 days after modeling, the treatment groups were administered different doses of magnolol (5, 10 and 20 mg/kg) by gavage every day, and a Morris water maze test was conducted after 2 months of treatment. The impacts of magnolol on amyloid β (Aβ) plaque deposition and neuroinflammation were assessed using Congo red and immunofluorescence staining. Immunofluorescence staining results were supplemented with western blotting and reverse transcription-quantitative PCR to ascertain the role of magnolol in other pivotal pathological mechanisms, including the formation of intracellular neurofibrillary tangles, compromised synaptic plasticity, and astrocyte and microglia activation. Administration of magnolol effectively mitigated cognitive impairment, reduced Aβ plaque deposition and inhibited neuroinflammation in Aβ1-42-induced mice. Moreover, hippocampal levels of tau, phosphorylated (p-)tau, glycogen synthase kinase 3β (GSK3β), p-GSK3β, synaptophysin, brain-derived neurotrophic factor, glial fibrillary acidic protein and ionized calcium binding adaptor molecule 1 revealed that magnolol also limited neurofibrillary tangle formation, repaired synaptic plasticity, and inhibited astrocyte and microglia activation. In conclusion, the present findings broaden the current understanding of the mechanisms explaining the neuroprotective effects of magnolol against AD progression. Notably, it may inhibit multiple manifestations of AD, including plaques and neuroinflammation, while also exhibiting the capacity to restore AD-related neurological damage.

Abstract 4138014: Incidence and Association of Amyloidosis Found at Carpal Tunnel Release to Development of Cardiac Involvement

Background: Carpal tunnel syndrome (CTS) associated with transthyretin cardiac amyloidosis (ATTR-CA) may manifest 5 to 10 years prior to cardiac diagnosis. Transthyretin amyloidosis (ATTR) identification in the tenosynovium isolated during carpal tunnel release (CTR) may facilitate earlier ATTR-CA diagnosis and treatment. Hypothesis: Amyloid discovered at CTR will be associated with, and/or serve as a precursor for, cardiac involvement. Aim: We aimed to determine the frequency and type of amyloid found at CTR as well as the incidence of cardiac involvement and/or treatment indication. Methods: A retrospective analysis was conducted on a convenience sample of adults undergoing CTR from 01/05/2021 to 03/24/2023. Biopsy specimens with confirmed amyloid via Congo red staining were analyzed using mass spectrometry (MS) for subtyping. Patients with amyloid in tenosynovial tissue were offered cardiac evaluation with history and physical, electrocardiogram, B-type natriuretic peptide, troponin-I HS, echocardiogram, and TcPYP nuclear scintigraphy. Patients with light-chain amyloidosis (AL) were referred to oncology, while patients with ATTR were offered genetic testing. Results: A total of 578 patients underwent CTR with tenosynovial sample excision. Amyloid involvement by Congo red staining was found in 43 patients (7.4%), with a definitive subtype confirmed by MS in 30 patients (5.2%). Samples were insufficient to define subtypes in 13 patients. Sole ATTR was found in 27 patients (4.7%), both AL and ATTR were found in 1 patient, and sole AL was found in 2 patients. Of the 28 ATTR patients (the patient with both ATTR and AL included), 24 pursued genetic testing, revealing all had wild-type ATTR. Of these 28 patients, 21 underwent cardiac evaluation, for which 2 had cardiac involvement on TcPYP and started tafamidis therapy. The patients with concomitant ATTR and AL or sole AL did not have evidence of cardiac involvement, however, chemotherapy was initiated as indicated. Unfortunately, 1 patient with AL on CTR did not pursue an evaluation. Conclusion: Incidental amyloid tissue found at CTR is common and associated with occult involvement warranting therapy in a minority of cases. Future cardiac involvement is unknown, and this cohort will be followed to determine future incidence. A startling number of patients were found to have occult systemic AL discovered at CTR. These incidental diagnoses may trigger lifesaving interventions, which may otherwise go undiagnosed.

Abstract 4112944: A diagnostic challenge overcome with persistent clinical suspicion in a case of cardiac AL amyloidosis

Case: A 62-year old male with a history of chronic kidney disease and paroxysmal atrial fibrillation presented with several months of fatigue, a progressive decline in functional status (NYHA IIIb-IV), weight loss, and intermittent lower extremity swelling. Blood pressure was 92/72 mmHg on presentation, and heart rate was 90 bpm. Physical examination revealed a jugular venous pressure of 12-14 cm H2O and 2+ bilateral lower extremity edema with mild wheezing. Methods/workup: Laboratories are shown in Table 1 and were notable for a serum free light chain ratio of 0.14. Transthoracic echocardiogram (figure 1) revealed a left ventricular (LV) ejection fraction of 33% with increased LV wall thickness and severe bi-atrial enlargement. Bone marrow biopsy revealed 2% lambda restricted plasma cells without evidence of amyloid deposits; metastatic bone survey was negative. A fat pad biopsy and endomyocardial biopsy stained negative for amyloid with Congo red. Cardiac MRI showed diffuse, circumferential subendocardial late gadolinium enhancement, most compatible with cardiac amyloidosis (figure 2). The patient also had a chest CT scan that demonstrated a few patchy nodular pulmonary infiltrates. Given his extensive negative workup, he was referred to pulmonary and endobronchial biopsy was positive for amyloid on Congo red staining (figure 3). Biopsy tissue typing revealed Lambda subtype AL amyloidosis. Management: The patient was started on Daratumumab, Cyclophosphamide, Bortezomib, and Dexamethasone by Hematology. Unfortunately, about a month after his diagnosis, he developed worsening heart failure despite treatment. He was admitted to an outside hospital with refractory cardiogenic shock and passed away during that admission. Discussion: The diagnosis of AL amyloidosis requires a high index of suspicion for prompt diagnosis and treatment. We present a case in which cardiac amyloidosis was suspected based on the patient’s clinical picture and cardiac imaging; however, multiple initial biopsies were negative. An expedited, multimodal and interdisciplinary diagnostic assessment is imperative as prognosis with chemotherapy is highly dependent on extent of AL disease, patient functional status, and presence of cardiac compromise. Elevated natriuretic peptides, troponins, autonomic dysfunction, and hypotension are negative prognostic features.

Abstract 4120573: Abrupt cardiac rupture of the patient with ATTR amyloidosis

A Japanese male in his 80s was admitted to our emergency department with chest pain. His prior history was syncope and ventricular tachycardia. Transthoracic echocardiography (TTE) revealed thickening of the basal part of the antero-septal wall and thinning of the basal part of the inferior wall. Coronary angiography (CAG), cardiac MRI and blood test revealed no specific abnormalities. The day before his emergency admission, he visited the cardiology outpatient department for a periodic examination of cardiac hypertrophy. ECG, chest X-ray, and TTE were unchanged from his prior examination. However, that night, he experienced chest pain and was transferred to our hospital. On arrival, his vital signs were critical, with a blood pressure of 70/59 mmHg. ECG indicated ST elevation in antero-lateral leads. TTE detected pericardial effusion, and contrast-enhanced CT identified two focal areas of non-enhancement in the myocardium and pericardial effusion without signs of aortic dissection. CAG revealed no significant stenosis or obstructive lesion. During coronary angiography, the patient deteriorated to asystole. Cardiopulmonary resuscitation and percutaneous pericardial drainage were initiated. Hemorrhagic pericardial effusion was aspirated. Immediate surgical hemostasis was undergone, revealing about 2 cm lacerations and hematoma in the anterior and inferior wall. After surgery, he complained of abdominal discomfort. Esophageal gastrointestinal endoscopy was undergone to investigate the abdominal discomfort, and he was diagnosed with early gastric cancer. Endoscopic submucosal dissection (ESD) was undergone, and histopathological examination revealed adenocarcinoma and eosinophilic amorphous materials deposited in the submucosal vessels. Congo red staining of the submucosal vessels revealed orange-red areas. A technetium-99m pyrophosphatase scintigraphy showed Grade 2 uptake in the heart. Transthyretin (TTR) staining was positive in the specimen from ESD, leading to a diagnosis of TTR amyloidosis. Genetic test for TTR was negative, which led to a diagnosis of wild-type TTR amyloidosis and tafamidis was started. In cases of acute pericardial effusion and/or cardiac rupture, it is imperative to consider cardiac amyloidosis as a potential etiology, in addition to myocardial infarction and aortic dissection, especially in patients with left ventricular wall thickening, neuropathy, or abnormal ECG, also known as “red flag”.

Adverse Effects of Aβ1-42 Oligomers: Impaired Contextual Memory and Altered Intrinsic Properties of CA1 Pyramidal Neurons.

Aβ1-42 (amyloid beta) oligomers, the major neurotoxic culprits in Alzheimer's disease, initiate early pathophysiological events, including neuronal hyperactivity, that underlie aberrant network activity and cognitive impairment. Although several synaptotoxic effects have been extensively studied, neuronal hyperexcitability, which may also contribute to cognitive deficits, is not fully understood. Here, we found several adverse effects of in vivo injection of Aβ1-42 oligomers on contextual memory and intrinsic properties of CA1 pyramidal neurons. Male rats underwent behavioral and electrophysiological studies 1 week after microinjections into the dorsal CA1 region, followed by histological analysis. After 1 week, Aβ1-42 oligomers impaired contextual learning without affecting basic physiological functions and triggered training-induced neuronal excitability. Furthermore, riluzole, a persistent sodium current (INaP) blocker, dose-dependently reduced Aβ1-42 oligomer-induced hyperexcitability. Congo red staining, which detects insoluble amyloid deposits, further identified labeling of CA1 pyramidal neurons while immunohistochemistry with lecanemab, which detects soluble Aβ oligomers, revealed immunoreactivity of both pyramidal and non-pyramidal cells in the target area. Therefore, our study suggests that a single injection of Aβ1-42 oligomers resulted in contextual memory deficits along with concomitant neuronal hyperexcitability and amyloid deposition in the CA1 region after 1 week.

Oral administration of butylated hydroxytoluene induces neuroprotection in a streptozotocin-induced rat Alzheimer’s disease model via inhibition of neuronal ferroptosis

BackgroundAlzheimer’s disease (AD) is the most common human neurodegenerative disorder worldwide. Owing to its chronic nature, our limited understanding of its pathophysiological mechanisms, and because of the lack of effective anti-AD drugs, AD represents a significant socio-economic challenge for all industrialized countries. Neuronal cell death is a key factor in AD pathogenesis and recent studies have suggested that neuronal ferroptosis may play a major patho-physiological role. Since ferroptosis involves free radical-mediated lipid peroxidation, we hypothesized that enteral administration of the radical scavenger butylated hydroxytoluene (BHT) might slow down or even prevent the development of AD-related symptoms in an in vivo animal AD model.Material and methodsTo test this hypothesis, we employed the rat model of streptozotocin-induced AD and administered butylated hydroxytoluene orally at a dose of 120 mg/kg body weight. Following BHT treatment, neuronal cell death was induced by bilateral stereotactic intraventricular injection of streptozotocin at a dose of 3.0 mg/kg body weight. Three weeks after surgery, we assessed the learning capabilities and the short-term memory of three experimental groups using the conventional y-maze test: (i) streptozotocin-treated rats (BHT pre-treatment), (ii) streptozotocin-treated rats (no BHT pre-treatment), (iii) sham-operated rats (BHT pre-treatment but no streptozotocin administration). After the y-maze test, the animals were sacrificed, hippocampal tissue was prepared and several biochemical (malonyl dialdehyde formation, glutathione homeostasis, gene expression patterns) and histochemical (Congo-red staining, Nissl staining, Perls staining) readout parameters were quantified.ResultsIntraventricular streptozotocin injection induced the development of AD-related symptoms, elevated the degree of lipid peroxidation and upregulated the expression of ferroptosis-related genes. Histochemical analysis indicated neuronal cell death and neuroinflammation, which were paralleled by aberrant intraneuronal iron deposition. The streptozotocin-induced alterations were significantly reduced and sometimes even abolished by oral BHT treatment.ConclusionOur data indicate that oral BHT treatment attenuated the development of AD-related symptoms in an in vivo rat model, most probably via inhibiting neuronal ferroptosis. These findings suggest that BHT might constitute a promising candidate as anti-AD drug. However, more work is needed to explore the potential applicability of BHT in other models of neurodegeneration and in additional ferroptosis-related disorders.

Challenges in the Diagnosis and Treatment of AL Amyloidosis in a Resource-Limited Setting: A Retrospective Analysis of Clinical Characteristics and Outcomes at a Newly Established Amyloid Centre in India

Background: AL (light-chain) amyloidosis, the common form of the complex spectrum of amyloidosis, poses significant diagnostic and therapeutic challenges even in well-resourced healthcare settings owing to non-specific symptoms, delayed diagnosis, limited therapeutic options and the need for specialised diagnostic techniques. In resource-limited settings such as India, these challenges are further compounded by constraints in healthcare infrastructure, limited access to advanced diagnostic tools and high costs of treatment. In 2023, our hospital established an Amyloid Centre (Amrita Amyloid Center) aimed at improving the diagnosis, treatment, and overall management of amyloidosis. This retrospective study analyses clinical data from the past two decades to identify the key challenges faced in the diagnosis and treatment of AL amyloidosis in a resource-limited setting. Methodology: Retrospective data was retrieved from in-house electronic records using keywords (amyloid, amyloidosis light chain, systemic, primary) from February 2002 to July 2024. All patients diagnosed during this period were included. Diagnosis was based on clinical, biochemical, cardiac imaging and histopathological criteria consistent with AL amyloidosis. Patient data included demographics, presenting symptoms, organ involved, diagnostic modalities and treatment regimens [chemotherapy, stem cell transplantation (SCT), supportive care] .Outcomes measured included time to diagnosis (TTD) (from symptom onset to confirmed diagnosis), diagnostic challenges , therapy outcomes and treatment-related challenges .Temporal trends and associations with time of diagnosis were examined for all variables using conditional density plots, linear regression model with a restricted cubic spline, Spearman rank correlation test and F-test. All statistical analyses were conducted using R version 4.3.0. Results : A total of 448 patient records were screened. The following were excluded- confirmatory report unavailable n=139(31.0%), biopsy proven congophilic amyloidosis but no further data available n=56 (12.5%), cutaneous amyloidosis (lichen, macular, amyloid angiopathy) n=84 (18.8%) and amyloidosis with other histologies (AA, ATTR, others)n=52(11.6%). Out of 117 patients with AL amyloid, 79 (67.5%) were males. Median age of the cohort was 67(40-94) years. Common presenting symptoms were dyspnoea on exertion (35, 29.9%), pedal edema (26, 22.2%), gastrointestinal(GI) symptoms (17,14.5%) and weakness of extremities (8, 06.4%). Serum free light chain assays confirmed light chain restriction . Biopsied tissues which demonstrated amyloid with apple green birefringence in Congo Red stain included renal n=26(22.2%), duodenal/rectal n=32(27.4%), fat pad n=19(16.2%), bone marrow n=15(12.3%) and other tissues n=10(8.5%). Median TTD was 5.5 (0-147) months. Most common organ-system involved was cardiac 40 (34.2%), followed by renal 34 ( 29.0%), Gastrointestinal 17(14.5%), liver 4(3.4%), skin 6 (5.1%), lungs and musculo-skeletal 1(0.9%) each. Median dFLC was 283 (51-6150)mg/dL. Unequivocal findings of amyloidosis were seen in echocardiogram n=30(25.6%) and cardiac magnetic resonance n=12(10.3%). Revised Mayo 2012 Staging revealed Stage I (n=37;31.6%), Stage II (n=33;28.2%), Stage III (n=15;13.5%), Stage IV (n=25;21.4%) and not classified (n=6;5.1%) patients. Amyloid typing by mass spectroscopy (MS) was done for 8 (6.4%) patients. Four (3.4%) patients died before treatment initiation. Induction therapy of the remaining 113 patients consisted of thalidomide n=16(14.2%), Mel-Pred n=15(13.27%), CyBorDex n=62(54.86%), BorLenDex n=13 (11.5%), pomalidomide n= 3 (2.6%). Autologous SCT was done in 6 (5.3%)cases. Daratumumab was used in 4 (3.5%) patients .The median follow up was 9.5 (0-150) months, with 21(17.9%) deaths registered [ most common: sudden cardiac death n=11(9.4%) followed by sepsis and AKI n=4(3.4% each)].Kaplan-Meier curves were used to predict the overall survival of patients. At the end of one year, the overall survival rate was 59.2%. Conclusion:This study highlights the diagnostic delays and limited access to advanced diagnostic and therapeutic options for AL amyloidosis in a resource-limited setting, underscoring the need for improved healthcare infrastructure and accessible, cost-effective treatments to enhance patient outcomes.

Early Onset Renal Amyloidosis in Section a Case of Rheumatoid Arthritis: A Case Report

Amyloidosis refers to a group of disorders characterised by the deposition of misfolded amyloid proteins in various tissues, leading to organ dysfunction. Secondary (AA) amyloidosis is most commonly associated with chronic inflammatory conditions such as Rheumatoid Arthritis (RA), where persistent inflammation triggers the overproduction of Serum Amyloid A (SAA) protein, which subsequently deposits as amyloid fibrils. Renal involvement is frequent in AA amyloidosis, often manifesting as nephrotic syndrome due to amyloid deposition in the glomeruli. Present case is of a 38-year-old female with a five-year history of RA, who was on methotrexate and sulfasalazine, and presented with frothy urine and progressive anasarca over two months. She exhibited a highgrade fever, cough, shortness of breath and right-sided chest pain. Examination revealed bilateral pedal oedema, facial puffiness, and signs of pleural effusion and ascites. Urine examination indicated nephrotic-range proteinuria, and pleural fluid analysis revealed a transudative effusion. A kidney biopsy confirmed AA amyloidosis, with positive Congo red staining and apple-green birefringence. This case highlights the unusual early onset of AA amyloidosis in RA, developing only five years postdiagnosis, significantly earlier than the typical median of 19 years. Despite well-managed RA and minimal disability, the patient developed nephrotic syndrome and subsequent immunosuppression, culminating in empyema. This case underscores the unpredictable progression of RA-related complications and highlights the critical need for vigilant monitoring and proactive management strategies.

Role of BIX01294 in the intracranial inhibition of H3K9 methylation lessens neuronal loss in vascular dementia model.

Dementia develops as a result of multiple factors, including cerebrovascular disease which is called vascular dementia (VD). Histone-3 lysine-9 dimethylation (H3K9me2) broadly increases during VD and inhibits neuroprotective gene expressions. So, we aimed to determine how H3K9me2 inhibitor (BIX01294) affects neuronal damage in VD. An in vivo model of VD was used followed by BIX01294 treatment. Behavioral tests, hematoxylin, and eosin (H&E), Congo red, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining were carried out. Hippocampal phosphorylated cyclic-AMP responsive element binding protein (p-CREB), c-fos, brain-derived neurotrophic factor (BDNF), and H3K9me2, were detected by western blot analysis technique. Neurological deficit and anxiety-related behavior significantly reduced in the treatment group compared to the VD group (p < 0.05). BIX01294 improved spatial and passive avoidance memory (p < 0.01 and p < 0.05, respectively) compared to the VD group. Treatment with BIX01294 restored the level of p-CREB/CREB ratio (p < 0.05), cfos (p < 0.01), BDNF (p < 0.01), and suppressed H3K9me2 (p < 0.001) when compared to the VD group. BIX01294 microinjection reduced the apoptosis level in TUNEL staining (p < 0.05), and raised neural cell count in H&E staining (p < 0.01); amyloid beta accumulation significantly decreased in the treatment group (p < 0.05) compared to the VD group. In conclusion, long-term treatment with a low dose of BIX01294 can prevent the progression of neuronal loss in VD model by raising the expression of neurotrophic factors, and reducing the apoptosis level.

Down regulation of the c-Fos/MAP kinase signaling pathway during learning memory processes coincides with low GnRH levels in aluminum chloride-induced Alzheimer's male rats.

Aluminum chloride (Al) is associated with Alzheimer's disease (AD) and reproductive disorders. But the relationship between gonadotropin-releasing hormone (GnRH) and c-Fos levels, the end product of MAP-kinase signaling, in AD is unknown, so we aimed to investigate this relationship. We exposed rats to Al dissolved in drinking water (10 and 50mg/kg) for two and four weeks. The control group received only drinking water. At the end, the blood sample was collected under deep anesthesia and the brain was dissected on ice, and the testicular tissue was fixed in formalin. Amyloid beta (βA) plaques in brain regions and the number of CA1 neurons were evaluated by Congo red staining and cresyl violet staining. Activation of neuronal nitric oxide synthase (nNOS) was studied using NADPH-diaphorase. The levels of c-Fos and testosterone receptors in the target area were examined immunohistochemically. Brain GnRH levels were determined by blotting, and serum levels of gonadotropins and steroids were measured by enzyme-linked immunosorbent assay (ELISA). All data were analyzed using analysis of variance (ANOVA) at α = 0.05 level. The accumulation of βA plaque was observed along with a decrease in the number of CA1 pyramidal neurons and a significant decrease in the levels of c-Fos and GnRH in the brains of rats receiving Al, which was aligned with a significant decrease in serum levels of testosterone and LH. This study, for the first time, showed a link between dementia and a concomitant decrease in brain GnRH and c-Fos levels.

Tumefactive Pulmonary amyloidosis in an elderly male: A Diagnostic Dilemma

Abstract Introduction/Objective Tumefactive pulmonary amyloidosis is a rare variant of pulmonary amyloidosis characterized by the extracellular deposition of abnormal protein fibrils in the lung parenchyma, forming tumor-like masses called amyloidomas. Although most cases are clinically silent, it may present with findings that mimic a wide range of respiratory conditions (eg. primary or metastatic malignant nodules, granulomatous disease, multiple pulmonary hamartomas etc), thereby necessitating a comprehensive and broad-based approach to differential diagnosis. Methods/Case Report We present a case of an 88-year-old male with a history of smoking and multiple comorbidities. He presented with numerous bilateral pulmonary nodules identified on a CT scan, concerning for malignancy. A biopsy from a left upper lobe lesion revealed necrotic tissue and was non-diagnostic. A PET scan to assess for metastatic disease was unremarkable, and demonstrated mild hypermetabolic activity, confined to the lungs, with standardized uptake values (SUV) ranging from 4.3 to a maximum SUV of 8.9. A subsequent right lung biopsy was positive for amyloid, confirmed by Congo red staining. A Mass spectrometry analysis further identified the amyloid as AL type. Serum, and urine protein electrophoresis with immunofixation were negative for M protein and the serum free light chain assay was normal as well. Significantly, the bone marrow biopsy also yielded normal results, and the patient was diagnosed with a localized form of AL amyloidosis. Results (if a Case Study enter NA) NA Conclusion This case highlights how tumefactive pulmonary amyloidosis poses a diagnostic challenge due to its rarity and resemblance to malignancy on imaging. Careful histopathological examination, including immunohistochemical stains and amyloid typing, was pivotal in establishing the correct diagnosis and underscores the essential role of pathology and laboratory medicine in differentiating rare pulmonary conditions, along with the need for a multidisciplinary approach in complex diagnostic scenarios.

Secondary Amyloidosis Treated with Tocilizumab as a Complication of Temporal Arteritis.

Temporal arteritis is a large-vessel vasculitis mostly seen in the elderly. Amyloidosis may be secondary to a chronic inflammation of body organs. Here, we present the second case report of temporal arteritis complicated by amyloidosis that was successfully treated by tocilizumab. A 64-year-old female presented complaining of fatigue, fever, and diarrhea accompanied by abdominal pain. One year before presentation, she was diagnosed with temporal arteritis. She was treated with 15 mg/day oral prednisone for the last 6 months, with partial remission, but persistence of the fatigue and an elevated erythrocyte sedimentation rate (ESR, 56 mm/h). Physical examination showed tenderness of both temporal arteries and a soft abdomen. Colon tissue biopsy showed amyloid depositions in the vessels and stroma that were positive for Congo red staining. Tocilizumab was started with 8 mg/kg intravenous, the diarrhea resolved, and the arthralgia improved within 1 month, with a decrease in the ESR to 8 mm/h, and a C-reactive protein (CRP) level of 0.98 mg/dl. Monthly tocilizumab therapy remains efficacious 12 months later and was stopped due to lack of tocilizumab from the hospital. No side effects of tocilizumab were registered. Chronic inflammation may be complicated by amyloidosis in patients with rheumatic diseases and genetic predisposition. Therefore, it is important to screen for intestinal Amyloid A (AA) amyloidosis in individuals with gastrointestinal disorders complicated by rheumatic disorders. AA amyloidosis may be complicated by temporal arteritis and presented with gastrointestinal symptoms such as diarrhea. Amyloidosis is manifested by the deposition of insoluble protein aggregates in organs.Amyloid A (AA) amyloidosis occurs as a complication of chronic inflammation in patients with a genetic predisposition to rheumatic diseases.Temporal arteritis complicated with AA amyloidosis is extremely rare.

Identification and Growth Characteristics of Cellulase Producing Bacteria

This study was envisaged to from fresh cow dung using the Congo red staining method, and screened by measuring their endoglucanase activity (CMC enzyme activity), exoglucanase activity (Cex enzyme activity), and filter paper enzyme activity (FPA enzyme activity). Subsequently, the molecular, morphological characteristics, and growth characteristics were studied. Three strains showed clear transparent circles on the Congo red cellulose medium. LY3 exhibited strong enzyme production ability, with relatively higher enzyme activity than the other two strains. LY3 was found to be a dominant strain that required a temperature of 40℃and pH 7.0 for optimal growth. Based on the molecular and morphological characteristics, the three strains were preliminarily identified as Bacillus. Bangladesh J. Bot. 53(3): 849-856, 2024 (September) Special

L’amylose – point de vue du pathologiste

Despite the advent of scintigraphic diagnosis of transthyretin amyloidosis, the role of pathologists remains central for the diagnosis and typing of amyloidosis. The anatomo-pathological diagnosis and typing of amyloidosis are hindered by pitfalls that should be known, linked to the Congo red staining (technical implementation and interpretation) and the difficulties of immunohistochemistry. The use of expert centers is recommended for difficult cases and when the type cannot be confirmed.

7289 An interesting case of Diabetes Mellitus Type 1 induced Renal Tubal Acidosis Type 1

Abstract Disclosure: R. Ripa: None. A. Itani: None. S.K. Patel: None. Introduction: Renal Tubular Acidosis (RTA) are a group of disorders that involve dysfunctional transporters and channels in the renal system. These disorders can be acquired (idiopathic, medication side effects, and medical disorders) or inherited. The most common type worldwide is type 4 RTA, the main cause is diabetic nephropathy leading to hypoaldosteronism. The association of type 1 RTA with diabetes has rarely been reported in studies. Here, we report the presentation, diagnosis and management of a rare case of type 1 RTA in a patient with a history of diabetes mellitus type 1. Case: A male in his 50s with a 30-year history of uncontrolled type 1 diabetes mellitus on insulin pump and coronary artery disease presented to his cardiologist with worsening chest pain, fatigue, polyuria, weight loss of 15 lbs, and recent insulin pump change. Blood tests revealed serum bicarbonate of 10 mmol/L and he was sent to the hospital. On examination, he was hypotensive and bradycardic with dry mouth and decreased skin turgor. His blood tests were consistent with type 1 RTA, he was treated with intravenous fluids containing bicarbonate plus potassium and transitioned to oral potassium citrate on discharge. He underwent renal biopsy as work up was unrevealing for a cause of type 1 RTA. Biopsy revealed normal glomeruli with negative immunofluorescence and Congo red stain. The biopsy, however, did show mild patchy chronic interstitial fibrosis with lymphocytes and mild tubular atrophy. At one year follow up, he remained on oral potassium citrate with resulting serum bicarbonate of 26 mmol/L and a normal renal function. Discussion: Diabetes mellitus can lead to a variety of renal pathologies, specifically metabolic acidosis that can be either seen as diabetic ketoacidosis or hyporeninemic hypoaldosteronism (Type 4 RTA). In our patient, despite being a diabetic, he did not present with either elevated anion gap, ketonuria or hyperkalemic hyperchloremic acidosis typical of type 4 RTA. He was noted to have hypokalemic hyperchloremic acidosis, with a positive urine anion gap, and an alkaline urine, all characteristic of type 1 RTA. Acquired distal RTA occurs more commonly due to certain medications or autoimmune diseases as in Sjögren’s syndrome. The relationship between distal RTA and Sjögren’s syndrome has already been established, unlike the relationship with type 1 diabetes. There have been few case reports of distal RTA in association with type 1 diabetes mellitus, implying a possible autoimmune pathology. Prior case reports also not only had patients who were insulin dependent, but diabetes was present for more than a decade at least suggesting chronicity of disease is needed. Traditional management consists of alkaline therapy and treatment of the underlying disease, or removal of offending medication. In this case, we hope to highlight distal RTA in a diabetic who doesn’t meet criteria for DKA or type 4 RTA. Presentation: 6/1/2024

Development of keratin-based fibers fabricated by interfacial polyelectrolyte complexation for suture applications

Interfacial Polyelectrolyte Complexation (IPC) is a convenient way to produce composite, micro-scale fibers. In this paper, we report the successful development of novel keratin-based IPC fibers and also demonstrate the feasibility of using these fibers as sutures through a proof-of-concept in vivo study. Two composite fibers were produced: chitosan-keratin (CK) and keratin-keratin (KK). These fibers were evaluated for their physico-chemical, mechanical and biochemical properties. In the dry state, the CK fiber had a greater Young's modulus of about 2 GPa while the KK fiber registered a longer strain-at-break of about 100 % due to the strain-stiffening effect. Notably, the keratins were found to assemble into amyloids within the composite fibers based on Congo red staining and Wide-Angle X-Ray Scattering. Functionally, both fibers were malleable could be weaved, braided and knotted. When used as sutures to close incisional wounds in mice over 21 days, these fibers were found to elicit minimal host tissue response and were partially degraded over the duration. Interestingly, the KK fiber evoked a lower extent of immune cell response and fibrous capsule encapsulation that was comparable to commercial, non-absorbable Dafilon® sutures. This work demonstrated the possibility of producing keratin-based IPC fibers which may find practicality as medical sutures.