Isolation of a galactose-free 20-kDa fragment exhibiting butyrylcholine esterase and aryl acylamidase activity from human serum butyrylcholine esterase by limited α-chymotrypsin digestion

Purified human serum butyrylcholine esterase (~ 90-kDa subunit), which also exhibits aryl acylamidase activity, was subjected to limited α-chymotrypsin digestion. Three major protein fragments of ~ 50 kDa, ~ 21 kDa and ~ 20 kDa were found to be produced, as observed by SDS-gel electrophoresis of the chymotryptic digest. The purified butyrylcholine esterase could fully bind to a Ricinus-communis-agglutinin-Sepharose column but after chymotryptic digestion about 15-20% of the enzyme activity remained unbound and was recovered in the run-through fractions. Sephadex G-75 chromatography of the chymotryptic digest showed an enzymatically active fragment eluted at an approximate molecular mass of 20 kDa, apart from the undigested butyrylcholine esterase eluted at the void volume. The butyrylcholine esterase fragment that did not bind to Ricinus communis agglutinin also was eluted at an approximate molecular mass of 20 kDa from a Sephadex G-75 column. This enzymatically active low-molecular-mass fragment from Sephadex G-75 chromatography showed a single protein band of ~ 20 kDa on SDS-gel electrophoresis. Neutral sugar analysis of the ~ 20 kDa fragment showed the presence of mannose only, whereas the undigested butyrylcholine esterase showed the presence of both mannose and galactose. Amino-terminal-sequence analysis of the ~ 20 kDa fragment showed the sequence Arg-Val-Gly-Ala-Leu, which agrees with amino acid residues 147-151 reported for human serum butyrylcholine esterase [Lockridge et al. (1987) J. Biol. Chem. 262, 549-557]. Both cholinesterase and aryl acylamidase activities were co-eluted in all chromatographic procedures. The results suggested that limited α-chymotrypsin digestion of human serum butyrylcholine esterase resulted in the formation of a ~ 20-kDa enzymatically active fragment with Arg147 as its N-terminal residue and which was devoid of galactose

Localization of the peptidase activity of human serum butyrylcholinesterase in a – 50-kDa fragment obtained by limited α-chymotrypsin digestion

Purified human serum butyrylcholinesterase (∼90-kDa subunit) is known to exhibit aryl acylamidase and peptidase activity. Limited α-chymotrypsin digestion of the purified butyrylcholinesterase gave three major protein fragments of ∼ 50 kDa, ∼21 kDa and ∼ 20 kDa. In our earlier studies [Rao and Balasubramanian (1989) Eur. J. Biochem. 179, 639-644] we characterized the ∼ 20-kDa fragment and showed that it exhibited both butyrylcholinesterase and aryl acylamidase activities. In the present studies the ∼ 50-kDa fragment is characterized. This fragment, after isolation by Sephadex G-75 chromatography from a chymotryptic digest of purified butyrylcholinesterase, exhibited only peptidase activity and was devoid of cholinesterase and aryl acylamidase activities. It could bind to a column of Ricinus communis agglutinin bound to Sepharose, indicating its glycosylated nature and the presence of galactose. The peptidase activity in the ∼50-kDa fragment could be immunoprecipitated by a polyclonal antibody raised against purified butyrylcholinesterase. SDS-gel electrophoresis of this fragment isolated by R. communis agglutinin - Sepharose and Sephadex G-75 chromatography showed a protein band of ~ 50 kDa by silver staining. Amino-terminal sequence analysis of the ∼ 50-kDa fragment gave the sequence of Gly-Pro-Thr-Val-Asp which corresponded to amino acid residues 291-295 in the butyrylcholinesterase sequence [Lockridge et al. (1987) J. Biol. Chem. 262, 549-557]. The combined results suggested that α -chymotrypsin digestion of human serum butyrylcholinesterase resulted in the formation of a ∼ 20-kDa fragment exhibiting both cholinesterase and aryl acylamidase activities and a ∼50-kDa fragment exhibiting only peptidase activity

Intrathecal baclofen in management of a patient with very severe tetanus

Tetanus is uncommon in developed countries. The majority of tetanus cases occur in third world countries and 50% of these cases occur in neonates. There are more than 800,000 deaths due to tetanus each year in the world. We present a case of 40-year-old male patient diagnosed to have very severe tetanus - Grade IV as per Ablett classification of severity, managed in our hospital with aggressive treatment for 27-days and use of intrathecal baclofen he showed drastic improvement in this status.  He was discharged in neurological intact conditions with hemodynamic stability.

The Entrapment of Unfree Labor: Theory and Examples from India

In this article we explore some aspects of contemporary unfree labor in rural south India. We draw on 130 case studies and (informally) extensive field research. We do so in order to make the central point that the conditions of unfreedom are variable and subject to change but that the basic vulnerabilities are significant. Being unfree in a labor relationship is a contingent effect of a set of factors. We stress the role of (a) entrapment of laborers, (b) immiseration within bondage, and (c) barriers to exit from the labor contract. In explanations, structural factors are also important. The article forms a basis for further empirical research in a variety of global settings even beyond India

Ocrelizumab versus Interferon Beta-1a in Relapsing Multiple Sclerosis

Supported by F. Hoffmann–La Roche

A local human Vδ1 T cell population is associated with survival in nonsmall-cell lung cancer

Murine tissues harbor signature γδ T cell compartments with profound yet differential impacts on carcinogenesis. Conversely, human tissue-resident γδ cells are less well defined. In the present study, we show that human lung tissues harbor a resident Vδ1 γδ T cell population. Moreover, we demonstrate that Vδ1 T cells with resident memory and effector memory phenotypes were enriched in lung tumors compared with nontumor lung tissues. Intratumoral Vδ1 T cells possessed stem-like features and were skewed toward cytolysis and helper T cell type 1 function, akin to intratumoral natural killer and CD8+ T cells considered beneficial to the patient. Indeed, ongoing remission post-surgery was significantly associated with the numbers of CD45RA−CD27− effector memory Vδ1 T cells in tumors and, most strikingly, with the numbers of CD103+ tissue-resident Vδ1 T cells in nonmalignant lung tissues. Our findings offer basic insights into human body surface immunology that collectively support integrating Vδ1 T cell biology into immunotherapeutic strategies for nonsmall cell lung cancer

Ayurveda and the science of aging

Since time immemorial, humanity has been concerned with developing and preserving youthful vigor, and extending longevity by stopping or delaying the aging process. By 2030, one in five of the world population will be over 65 years old. Longevity and old age are accompanied with a variety of health challenges and population studies indicate that the elderly will use between three to five times more healthcare services compared to the younger population. Modern medicine has made a great deal of progress in understanding the aging process and in controlling age-associated health issues including heart attacks, strokes, diabetes, cancer, senility, and arthritis. Thus, every individual is now looking forward to a youthful, productive lifespan of 100 or more years filled with unlimited health and opportunity. Research by aging experts is focused on ways to go against the natural order of the aging process in order to delay it. Interventions include among other things anti-aging pills, restricted food consumption and cloning body parts to stay young and delay biological aging. Ayurveda, one of the world's most authoritative mind-body-spirit medicinal systems, offers various concepts of the aging process. This system of medicine includes therapies for healthy aging so as to create an optimal health and lengthen an individual's healthspan by living in harmony with nature. This review will explore various aspects of aging and longevity by comparing the science of aging as defined by modern medicine with the Ayurvedic treatise of Jara and Vriddhavastha. Keywords: Aging, Lifespan, Healthspan, Ayurveda, Jara, Vriddhavastha, Parinam

Antiviral and Immunomodulation Effects of Artemisia

Background and Objectives: Artemisia is one of the most widely distributed genera of the family Astraceae with more than 500 diverse species growing mainly in the temperate zones of Europe, Asia and North America. The plant is used in Chinese and Ayurvedic systems of medicine for its antiviral, antifungal, antimicrobial, insecticidal, hepatoprotective and neuroprotective properties. Research based studies point to Artemisia’s role in addressing an entire gamut of physiological imbalances through a unique combination of pharmacological actions. Terpenoids, flavonoids, coumarins, caffeoylquinic acids, sterols and acetylenes are some of the major phytochemicals of the genus. Notable among the phytochemicals is artemisinin and its derivatives (ARTs) that represent a new class of recommended drugs due to the emergence of bacteria and parasites that are resistant to quinoline drugs. This manuscript aims to systematically review recent studies that have investigated artemisinin and its derivatives not only for their potent antiviral actions but also their utility against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Materials and Methods: PubMed Central, Scopus and Google scholar databases of published articles were collected and abstracts were reviewed for relevance to the subject matter. Conclusions: The unprecedented impact that artemisinin had on public health and drug discovery research led the Nobel Committee to award the Nobel Prize in Physiology or Medicine in 2015 to the discoverers of artemisinin. Thus, it is clear that Artemisia’s importance in indigenous medicinal systems and drug discovery systems holds great potential for further investigation into its biological activities, especially its role in viral infection and inflammation

The peptidase activity of human serum butyrylcholinesterase: studies using monoclonal antibodies and characterization of the peptidase

Purified human serum butyrylcholinesterase, which exhibits cholinesterase, aryl acylamidase, and peptidase activities, was cross-reacted with two different monoclonal antibodies raised against human serum butyrylcholinesterase. All three activities were immunoprecipitable at different dilutions of the two monoclonal antibodies. At the highest concentration of the antibodies used, nearly 100% of all three activities were precipitated, and could be recovered to 90-95% in the immunoprecipitate. The peptidase activity exhibited by the purified butyryl-cholinesterase was further characterized using both Phe-Leu and Leu-enkephalin as substrates. ThepH optimum of the peptidase was in the range of 7.5-9.5 and the divalent cations Co2+, Mn2+, and Zn2+ stimulated its activity. EDTA and other metal complexing agents inhibited its activity. Thiol agents and -SH group modifiers had no effect. The serine protease inhibitors, diisopropylfluorophosphate and phenyl methyl sulfonyl fluoride, did not inhibit. When histidine residues in the enzyme were modified by diethylpyrocarbonate, the peptidase activity was not affected, but the stimulatory effect of Co2+, Mn2+, and Zn2+ disappeared, suggesting the involvement of histidine residues in metal ion binding. These general characteristics of the peptidase activity were also exhibited by a 50 kD fragment obtained by limited α-chymotrypsin digestion of purified butyrylcholinesterase. Under all assay conditions, the peptidase released the two amino acids, leucine and phenylalanine, from the carboxy terminus of Leu-enkephalin as verified by paper chromatography and HPLC analysis. The results suggested that the peptidase behaved like a serine, cysteine, thiol-independent metallopeptidase