Bioactive potential of Lagotis cashmeriana: a study on morphology, phytochemicals, and antioxidant activity.

The present study aimed to investigate the morphological features, phytochemicals, phenolic content, and antioxidant activity in different parts of Lagotis cashmeriana. The morphological features depicted that the plant is 7.9 ± 1.699 cm tall with flowers arranged into an inflorescence. The length of inflorescence was 2.597 ± 0.796 cm. Basal leaves were measuring 2.99 ± 0.58 cm. Besides, the number of basal leaves and inflorescence ranged from 4-9 and 0-4 respectively. Methanolic extract of leaves displayed the highest phenolic content (169.5 µg/mL of GAE), followed by inflorescences (157 µg/mL of GAE). Among aqueous extracts, leaves displayed the highest phenolic content (88.38 µg/mL of GAE), followed by inflorescences (76.95 µg/mL of GAE). The results of antioxidant study revealed that the methanolic extracts of leaves possessed the highest antioxidant potential (180.76 µg/mL of AAE). Interestingly, for each extract, there was a positive correlation between the phenolic content and the antioxidant activity.

Foreign body esophagus in a young infant.

Unattended children, mostly from low-socioeconomic contexts, who present with sudden onset obstructive respiratory and/or gastrointestinal symptoms, should be at high suspicion for foreign body ingestion. Prompt diagnosis helps avoid mismanagement and can potentially avoid dire outcomes.

Fetuin is the key for nanon self-propagation.

"Nanobacteria", also known as nanons or calciprotein particles (CPP), are nano-sized protein mineral complexes which have been isolated from numerous biological sources. Nanons possess self-replication properties and contain only serum proteins (e.g. Fetuin-A, Albumin). Herein, we develop a simplified in vitro model of nanons propagation composed of only fetuin-A as a protein. Using this model, we demonstrate that fetuin from nanons possesses a different, non-native conformation. Moreover, we show that nanons induce soluble fetuin-A precipitation which could serve as a template for calcification. This phenomenon explains the observed self-propagating properties that mimic infectious behavior. We also demonstrate that renal calculi are capable of inducing a conformational change in fetuin-A, suggesting that the propagation phenomenon of nanons may occur in vivo.

Isolation, propagation, and analysis of biological nanoparticles.

Calcifying biologic nanoparticles (NPs) have been implicated as nucleation points for a number of -pathologic events that include vascular calcification and the formation of kidney stones. In order to study these potential relationships, reproducible isolation of well-characterized biologic NPs is a necessity. Our group has isolated and propagated calcifying NPs from several human tissues and renal stones. Specific proteins that could nucleate a calcium phosphate shell under physiologic conditions have been identified as part of their structure, including elongation factor Tu (EF-Tu) and fetuin-A. Visualization, using advanced transmission electron microscopy (TEM), immunofluorescence microscopy, and nuclear and antibody staining in conjunction with flow cytometry, can further elucidate NPs composition and their role in pathology. In order to allow uniform investigation by others, the isolation, culture, and handling procedures for biologic NPs from human calcified vascular tissue and kidney stones are reported in detail.

Key role of alkaline phosphatase in the development of human-derived nanoparticles in vitro.

Alkaline phosphatase (ALP) is an enzyme critical for physiological and pathological biomineralization. Experiments were designed to determine whether ALP participates in the formation of calcifying nanometer sized particles (NPs) in vitro. Filtered homogenates of human calcified carotid artery, aorta and kidney stones were inoculated into cell culture medium containing 10% fetal bovine serum in the absence or presence of inhibitors of ALP or pyrophosphate. A calcific NP biofilm developed within 1 week after inoculation and their development was reduced by pyrophosphate and inhibitors of ALP. ALP protein and enzymatic activity were detected in washed NPs, whether calcified or decalcified. Therefore, ALP activity is required for the formation of calcifying NPs in vitro, as has previously been implicated during pathological calcification in vivo.

Proteomic evaluation of biological nanoparticles isolated from human kidney stones and calcified arteries.

Calcifying biological nanoparticles (NPs) develop under cell culture conditions from homogenates of diverse tissue samples displaying extraosseous mineralization, including kidney stones and calcified aneurysms. Probes to definitively identify NPs in biological systems are lacking. Therefore, the aim of this study was to begin to establish a proteomic biosignature of NPs in order to facilitate more definitive investigation of their contribution to disease. Biological NPs derived from human kidney stones and calcified aneurysms were completely decalcified by overnight treatment with ethylenediaminetetraacetic acid or brief incubation in HCl, as evidenced by lack of a calcium shell and of Alizarin Red S staining, by transmission electron microscopy and confocal microscopy, respectively. Decalcified NPs contained numerous proteins, including some from bovine serum and others of prokaryotic origin. Most prominent of the latter group was EF-Tu, which appeared to be identical to EF-Tu from Staphylococcus epidermidis. A monoclonal antibody against human EF-Tu recognized a protein in Western blots of total NP lysate, as well as in intact NPs by immunofluorescence and immunogold EM. Approximately 8% of NPs were quantitatively recognized by the antibody using flow cytometry. Therefore, we have defined methods to reproducibly decalcify biological NPs, and identified key components of their proteome. These elements, including EF-Tu, can be used as biomarkers to further define the processes that mediate propagation of biological NPs and their contribution to disease.

Lithogenesis: induction of renal calcifications by nanobacteria.

Nanobacteria have been isolated from kidney stones and it has been suggested that they may act as a nucleus for the initiation of the renal stones. In the present study, we examine their role in biocrystallization and their in vivo effects on kidney pathology. Calcium oxalate monohydrate (COM) assay was carried out in the presence of nanobacteria to study biocrystallization. Wistar rats were given an intravenous injection of nanobacteria and the kidneys were examined for pathological changes. The COM assay showed accelerated biocrystallization of (14)C-oxalate in the presence of nanobacteria, indicating them to be efficient candidates for biomineralization. Histopathological studies revealed bacteria induced renal tubular calcifications and various manifestations of infection. Our studies confirm that nanobacteria may be involved in the pathogenesis of renal tubular calcification.

Morphological and immunological characteristics of nanobacteria from human renal stones of a north Indian population.

The aim of this study was to detect, isolate and characterize the nanobacteria from human renal stones from a north Indian population, and to determine their role in biomineralization. Renal stones retrieved from the kidneys of 65 patients were processed and subjected to mammalian cell culture conditions. The isolated bacteria were examined using scanning (SEM) and transmission electron microscopy (TEM). They were characterized for the presence of DNA, proteins and antigenicity. The role of these bacteria in biomineralization was studied by using the (14)C-oxalate based calcium oxalate monohydrate (COM) crystallization assay. We observed the presence of apatite forming, ultrafilterable gram negative, coccoid microorganisms in 62% of the renal stones. SEM studies revealed 60-200 nm sized organisms with a distinct cell wall and a capsule. TEM images showed needle like apatite structures both within and surrounding them. They were heat sensitive, showed antibiotic resistance and accelerated COM crystallization. A potent signal corresponding to the presence of DNA was observed in demineralized nanobacterial cells by flow cytometry. The protein profile showed the presence of several peptide bands of which those of 18 kDa and 39kDa were prominent. Apatite forming nanosized bacteria are present in human renal stones and may play a role in the pathophysiology of renal stone formation by facilitating crystallization and biomineralization. However, further studies are required to establish the exact mechanism by which nanobacteria are involved in the causation of renal stones.