Complete genome sequence and transcriptome response to vitamin C supplementation of Novacetimonas hansenii SI1 - producer of highly-stretchable cellulose.

Novacetimonas hansenii SI1, previously known as Komagataeibacter hansenii, produces bacterial nanocellulose (BNC) with unique ability to stretch. The addition of vitamin C in the culture medium increases the porosity of the membranes and their stretchability making them highly moldable. To better understand the genetic background of this strain, we obtained its complete genome sequence using a hybrid sequencing and assembly strategy. We described the functional regions in the genome which are important for the synthesis of BNC and acetan-like II polymer. We next investigated the effect of 1% vitamin C supplementation on the global gene expression profile using RNA sequencing. Our transcriptomic readouts imply that vitamin C functions mainly as a reducing agent. We found that the changes in cellular redox status are balanced by strong repression of the sulfur assimilation pathway. Moreover, in the reduced conditions, glucose oxidation is decreased and alternative pathways for energy generation, such as acetate accumulation, are activated. The presence of vitamin C negatively influences acetan-like II polymer biosynthesis, which may explain the lowered yield and changed mechanical properties of BNC. The results of this study enrich the functional characteristics of the genomes of the efficient producers of the N. hansenii species. Improved understanding of the adaptation to the presence of vitamin C at the molecular level has important guiding significance for influencing the biosynthesis of BNC and its morphology.

Serum level of Selenium, Zinc, and Vitamin C and their relation to the clinical spectrum of leprosy.

INTRODUCTION: Leprosy is a chronic infectious disease with many risk factors including inadequate nutrient intake and nutritional deficiencies, which affect the immune system, and influence leprosy progression. OBJECTIVES: To elucidate the relation between the serum level of zinc, vitamin C, and selenium and the clinical spectrum of leprosy. METHODOLOGY: A case control study included 100 leprotic patients (50 multibacillary and 50 paucibacillary) and 100 age and sex matched controls. Vitamin C was measured by ELISA, zinc was measured by using centronic colorimetric spectrophotometry, and selenium was measured by Inductivity Coupled Plasma Optical Emission Spectroscopy technique. RESULTS: Zinc and Vitamin C levels were significantly lower in paucibacillary (mean ± SD = 89.86 ± 20.712 and 2.52 ± 1.27 respectively) and multibacillary (mean ± SD = 81.41 ± 18.61 and 1.98 ± 0.59 respectively) than in controls (mean ± SD = 107.34 ± 3.98 and 4.95 ± 2.45 respectively) (p value < 0.001) with no significant difference between paucibacillary and multibacillary patients (p value = 0.142 and = 0.066 respectively). Selenium level showed no significant difference between the three groups (p value > 0.05) (mean ± SD = 51.27 ± 42.61 in paucibacillary, 47.54 ± 30.21 in multibacillary, and 44.07 ± 46.58 in controls). CONCLUSIONS: Lower serum levels of zinc and vitamin C in leprosy patients may be a result of disease pathogenesis or related to the antioxidants based treatment. It might also present prior to the disease onset due to malnutrition that may have accelerated the development of leprosy.

M. leprae HSP18 suppresses copper (II) mediated ROS generation: Effect of redox stress on its structure and function.

Mycobacterium leprae, causative organism of leprosy, is known to counter redox stress generated by reactive oxygen species (ROS) during its survival inside host macrophages. But, the involvement of any antigenic protein(s) for countering such redox stress is still unknown. Interestingly, M. leprae HSP18, an important antigenic protein that helps in the growth and survival of M. leprae pathogen inside host macrophages, is induced under redox stress. Moreover, HSP18 also interacts with Cu2+. Copper (II) can induce redox stress via Fenton reaction. But, whether HSP18 suppresses Cu2+ mediated ROS generation, is still far from clear. Also, the effect of redox stress on its structure and function is not known. In this study, we show that HSP18 efficiently suppresses Cu2+ mediated generation of ROS and also prevents the redox mediated aggregation of a client protein (γD-crystallin). Upon exposure to substantial redox stress, irreversible perturbation in the secondary and tertiary structure of HSP18 and the tryptophan and tyrosine oxidation are evidenced. Interestingly, HSP18 retains a considerable amount of functionality even after being exposed to substantial redox stress. Perhaps, the redox scavenging ability as well as the chaperone function of HSP18 may possibly help M. leprae pathogen to counter redox stress inside host macrophages.

The efficacy of topical human amniotic membrane-mesenchymal stem cell-conditioned medium (hAMMSC-CM) and a mixture of topical hAMMSC-CM + vitamin C and hAMMSC-CM + vitamin E on chronic plantar ulcers in leprosy:a randomized control trial.

BACKGROUND: Healing of chronic plantar ulcers in leprosy (CPUL) typically takes a long time due to impaired neurological function, thereby reducing the levels of growth factors and cytokines. Cytokines can be found in metabolite products from amniotic membrane stem cells. Chronic ulcers are frequently characterized by high levels of reactive oxygen species. Vitamin E (α-tocopherol) is widely used in skin lesions, owing to its antioxidant and anti-inflammatory properties. Vitamin C also has antioxidant, anti-inflammatory, and collagen synthesis properties which are useful in wound healing. Herein, we compared the effects of topical human amniotic membrane-mesenchymal stem cell-conditioned medium (hAMMSC-CM) alone and with vitamins C and E on healing of CPUL. METHODS: In this randomized controlled trial, topical agents were applied every 3 days for up to 8 weeks. Ulcer size, side-effects, and possible complications were monitored weekly. RESULTS: Healing percentage increased each week in all groups. Mean difference in ulcer size was highest in the hAMMSC-CM + vitamin E group, implying better progress of wound healing. There were no side-effects or complications. CONCLUSIONS: hAMMSC-CM + vitamin E is best for healing of CPUL.

Combined vitamin C sonophoresis and neodymium-doped yttrium aluminum garnet (NdYAG) laser for facial hyperpigmentation: An outcome observation study in Asian patients.

BACKGROUND: The neodymium-doped yttrium aluminum garnet (NdYAG) laser therapy has been a popular technique for facial rejuvenation but certain adverse effects like post-inflammatory hyperpigmentation are issues of concern to Asian patients. AIMS: To assess the outcome following combined treatment with vitamin C sonophoresis and NdYAG laser, in selected cases of facial hyperpigmentation. METHODS: Twenty three women with dyschromia or melasma who had undergone five sessions of Q-switched NdYAG laser therapy followed by transdermal delivery of vitamin C via sonophoresis were selected after a retrospective review of case records. The objective and subjective clinical outcomes and the side effects, including erythema, scaling, pruritus, dryness and post-inflammatory hyperpigmentation were evaluated. RESULTS: In both objective or subjective outcomes, 91.3% (21/23) of the patients showed an excellent or better outcome, while 8.7% (2/23) showed no change. A majority of the patients (73.9%, 17/23) experienced no post-inflammatory hyperpigmentation or had slight post-inflammatory hyperpigmentation which quickly resolved within 1 week. Only one (4.3%) patient had extreme post-inflammatory hyperpigmentation which lasted for over a month. LIMITATIONS: This was a retrospective study without a control group; a comparative study with a control group (patients treated with the laser alone, without vitamin C sonopheresis) is needed to determine the difference in the outcome. CONCLUSION: The use of vitamin C sonophoresis along with NdYAG laser may reduce the incidence of adverse effects in Asian patients. Patients experienced obvious improvement in hyperpigmentation and had lower chances of experiencing extreme or severe post-inflammatory hyperpigmentation.

Saliva as a Diagnostic Tool for Measurement of Total Antioxidant Capacity in Children with Leprosy and Born to Leprosy Parent.

The aim of the study is to assess the role of saliva as a diagnostic tool for measurement of total antioxidant capacity in children with leprosy and children born to leprosy parent. One hundred fifty children in the age group of 4-15 years were split into three equal groups: children with leprosy (CL) and children born to leprotic parents (CLP) and healthy children. Vitamin C level was measured in saliva of children spectrophotometrically at 695nm by Phosphomolybdenum method. Data were determined with student's unpaired t test and one way ANOVA. The result of the study showed that children with leprosy exhibited significantly decreased salivary total antioxidant capacity as compared to healthy controls. Antioxidant Vitamin C was higher in the Paucibacillary leprosy (PB) than those of Multibacillary type (MB) (P < 0.001). As age advanced, there was a gradual increase in total antioxidant capacity in both the control and study groups and the results were highly significant statistically. Saliva is an easy medium.

Micronutrients influencing the immune response in leprosy / Micronutrientes que influyen en la respuesta immune en la lepra

La lepra es una enfermedad infecciosa crónica causada por el Mycobacterium leprae, un bacilo intracelular de transmisión aérea. La enfermedad afecta la piel y los nervios periféricos y causa secuelas neurológicas. El bacilo se multiplica lentamente en el hospedador y posiblemente la enfermedad ocurre por el mal funcionamiento de la respuesta inmunitaria del hospedador. Esta revisión aborda el papel de algunos micronutrientes específicos en la respuesta inmunitaria, tales como las vitaminas A, D, E, C, el cinc y el selenio, detallando sus mecanismos de acción en las enfermedades infecciosas y en la lepra. La respuesta inmunitaria a los patógenos libera sustancias nocivas que producen lesión tisular. Esta revisión también aborda cómo una menor cantidad de antioxidantes puede contribuir a un aumento del estrés oxidativo y a complicaciones de las enfermedades infecciosas y la lepra. Puesto que los micronutrientes poseen un efecto regulador de la respuesta inmunitaria innata y adaptativa, es importante un equilibrio perfecto de sus concentraciones para mejorar la respuesta inmunitaria frente a los patógenos (AU)

Leprosy is a chronic infectious disease caused by Mycobacterium leprae, an intracellular bacillus of airborne transmission. The disease affects the skin and peripheral nerves and can cause neurological sequelae. The bacillusmultiplies slowly in the host and the disease probably occurs due to malfunctioning in host immune response. This review addresses the role of some specific micronutrients in the immune response, such as Vitamins A, D, E, C, Zinc and Selenium, detailing their mechanisms of actions in infectious diseases, and in leprosy. The immune response to pathogens releases harmful substances, which lead to tissue damage. This review discusses how a decreased level of antioxidants may contribute to an increased oxidative stress and complications of infectious diseases and leprosy. As the nutrients have a regulatory effect in the innate and adaptative immune responses, a perfect balance in their concentrations is important to improve the immune response against the pathogens (AU)

Improvement of flow and bulk density of pharmaceutical powders using surface modification.

Improvement in flow and bulk density, the two most important properties that determine the ease with which pharmaceutical powders can be handled, stored and processed, is done through surface modification. A limited design of experiment was conducted to establish a standardized dry coating procedure that limits the extent of powder attrition, while providing the most consistent improvement in angle of repose (AOR). The magnetically assisted impaction coating (MAIC) was considered as a model dry-coater for pharmaceutical powders; ibuprofen, acetaminophen, and ascorbic acid. Dry coated drug powders were characterized by AOR, particle size as a function of dispersion pressure, particle size distribution, conditioned bulk density (CBD), Carr index (CI), flow function coefficient (FFC), cohesion coefficient using different instruments, including a shear cell in the Freeman FT4 powder rheometer, and Hansen flowability index. Substantial improvement was observed in all the measured properties after dry coating relative to the uncoated powders, such that each powder moved from a poorer to a better flow classification and showed improved dispersion. The material intrinsic property such as cohesion, plotted as a function of particle size, gave a trend similar to those of bulk flow properties, AOR and CI. Property improvement is also illustrated in a phase map of inverse cohesion (or FFC) as a function of bulk density, which also indicated a significant positive shift due to dry coating. It is hoped that such phase maps are useful in manufacturing decisions regarding the need for dry coating, which will allow moving from wet granulation to roller compaction or to direct compression based formulations.

From darkening urine to early diagnosis of alkaptonuria.

Alkaptonuria is a rare disorder of metabolism characterized by deficiency of homogentisic acid oxidase. Characteristic features include darkening of urine, ochronosis, and arthropathy. Darkening of urine is the only sign of the disorder in the pediatric age group, and it occurs at very early stage of the disorder, as reported by the parents. A 4-year-old boy presented to our clinic with the complaint of dark urine and bluish black staining of clothes. This darkening pointed to a positive physical history of bluish discoloration of sclerae which occurred off and on. We initiated treatment with ascorbic acid and a protein diet with restriction of phenylalanine and tyrosine (1.6 g/kg/d). This case report is significant because of the early diagnosis made.

Early detection of alkaptonuria.

Alkaptonuria is a rare disorder of metabolism characterized by deficiency of homogentisic acid oxidase. This leads to the characteristic features like darkening of urine, ochronosis and arthropathy. Darkening of urine is one of the first symptoms noticed by the parents of the child suffering from this disorder. Ochronosis is seen in various organs like eyes, skin, tendons and joints. A case of 10 year old boy is reported who was brought to this clinic with the presenting complaint of bluish discoloration of sclerae. This discoloration led to eliciting positive history of dark urine off and on. Further investigations confirmed alkaptonuria.

Assessment of antioxidant activity of some antileprotic drugs.

DPPH (alpha,alpha-diphenyl-beta-picryl hydrazyl) (CAS 217-591-8), a stable free radical can be used to determine the antioxidant activity (AOA) of some drugs. In the present study the DPPH method was used for the first time to test AOA of dapsone (CAS 80-08-0), clofazimine (CAS 2030-63-9) and rifampicin (CAS 13282-42-1) in vitro and deproteinated blood method. Ascorbic acid (CAS 50-81-7) was used as a control in the study, which showed concentration-dependent antioxidant activity. Rifampicin showed a per se effect but it showed concentration dependent decrease in the DPPH absorbance. Ascorbic acid, dapsone and rifampicin showed DPPH scavenging activity both in vitro and deproteinated blood method. Clofazimine did not have any influence on DPPH. This method may be extended to different drugs for testing their AOA in biological fluids.

Reduction of clastogenic effect of clofazimine, an antileprosy drug, by vitamin A and vitamin C in bone marrow cells of mice.

Clofazimine (CLF), an antileprosy drug, has earlier been proved to be clastogenic in mice in vivo. It is an important constituent of the triple-drug regimen recommended by WHO for the treatment of leprosy. In this study the protective role of vitamins A and C (vit A and vit C) against the clastogenic effect of CLF in mouse bone marrow cells has been evaluated. Two doses (20 and 40 mg/kg) of vit C and two doses (2500 and 5000 IU/kg) of vit A were tested against a dose of 40 mg CLF/kg. The drug alone induced chromosomal aberrations of about 8 times the control value. Neither of the doses of vit C exhibited any clastogenic effect and, when administered simultaneously with CLF, both reduced the effect of CLF very significantly, the higher dose reducing chromosomal aberrations almost to the control value. Conversely, both doses of vit A, when administered alone, brought about significant increases in chromosome aberrations over the control value; the higher, but not the lower dose, given simultaneously with CLF, minimized the effect of CLF significantly but not as greatly as vit C. A scavenging effect of the vitamins, removing free radicals produced by CLF, is assumed to be responsible for modulation of the clastogenic effect of CLF.

Energy generation mechanisms in the in vitro-grown Mycobacterium lepraemurium.

Mycobacterium lepraemurium was cultivated on Ogawa egg-yolk medium and its energy coupling mechanisms were investigated. Cell-free extracts prepared from in vitro-grown cells catalyzed phosphorylation coupled to the oxidation of generated NADH, added NADH, and succinate-yielding ratios of phosphorus moles incorporated into high-energy bonds to oxygen atoms utilized (P/O ratios) of 0.75, 0.52, and 0.36, respectively. Ascorbate oxidation alone or in the presence of tetramethyl-p-phenyline-diamine (TMPD) did not yield any adenosine triphosphate (ATP). However, ascorbate in the presence of added cytochrome c was coupled to ATP synthesis and yielded a P/O ratio of 0.12. The oxidative phosphorylation was uncoupled by all of the uncouplers used without any inhibition of oxygen consumption. ATP generation coupled to NADH oxidation was completely inhibited by the flavoprotein inhibitors, such as rotenone and amytal; these inhibitors had no effect, however, on ATP synthesis associated with succinate oxidation. Antimycin A or 2-n-heptyl-4-hydroxy-quinoline-N-oxide (HQNO) and cyanide inhibited markedly the oxidations of NADH and succinate as well as the coupled ATP generation. The phosphorylation coupled to ascorbate plus cytochrome c was not affected by either of the flavoprotein inhibitors or by antimycin A or HQNO, but was completely inhibited by cyanide. The thiol-bearing agents p-chloromercuribenzoate (PCMB) and N-ethylmaleimide were the potent inhibitors of the phosphorylation associated with the oxidation of NADH and succinate. The results indicate that the three energy-coupling sites are functional in the respiratory chain of in vitro-grown M. lepraemurium.

Comentários acerca do tratamento da hanseníase / Annotations on the therapy of leprosy

É importante na terapêutica da hanseníase näo serem olvidados certos princípios básicos de bioquímica e farmacologia que näo obedecidos, implicaräo em prejuízos para o tratamento. O presente trabalho focaliza três desses princípios: 1§ a sulfona e os corpos férricos têm açöes antagônicas frente ao M leprae e näo devem ser associados no tratamento da hanseníase; 2§ a DDS, como todo composto sulfamídico, tem açäo antivitamina B e seu uso, indiscriminado agrava os sintomas neurológicos da hanseníase; 3§ deve-se associar alcalino no tratamento da reaçäo hansênica, tendo em vista que este estado agudo favorece a acidose

A study of blood ascorbic acid in leprosy.

The basal level of blood ascorbic acid was studied in 70 cases of polar leprosy. The level was found to be significantly reduced in both polar types, more so in tuberculoid. In untreated cases of these two polar forms, the differences were highly significant. No significant differences were found between untreated and treated cases of either form of leprosy, indicating that dapsone (DDS) has no effect on the ascorbic acid level. In lepromatous leprosy, the level was not affected by erythema nodosum leprosum (ENL). The deficiency of ascorbic acid in leprosy might be due to the disease per se and/or associated with nutritional factors. In trophic ulcers after supplement therapy for 60 days, the low level of ascorbic acid became close to the control group with marked improvement in the healing of the ulcers clinically. In addition, lactic and pyruvic acids were also measured and found to be significantly raised. After ascorbic acid supplement, the levels fell to near the control values, indicating that ascorbic acid played a role in controlling the infection as well as possible thiamine deficiency and also checked the disturbance caused by DDS on lactate and pyruvate metabolism. The present study suggested that supplementing DDS therapy with ascorbic acid might provide a beneficial effect on the general condition of the leprosy patients and the early healing of trophic ulcers.

Effects of ascorbate on normal and abnormal leucocyte functions.

The stimulatory effects of ascorbate on neutrophil motility in vitro and in vivo and lymphocyte transformation to mitogens following ingestion or intravenous injection of ascorbate have been found to be related entirely to inhibition of the autooxidative effect of the myeloperoxidase/hydrogen peroxide/halide system (MPO/H2O2/halide system). Stimulation of neutrophil migration and lymphocyte transformation following a single intravenous injection of 1 g of ascorbate was associated with inhibition of the MPO/H2O2/halide system. The immunostimulatory activity and peroxidase inhibitory activity was related entirely to the serum ascorbate level. The relationship between inhibition of the peroxidase/h2O2/halide system and stimulation of neutrophil motility and lymphocyte mitogen-induced transformation was further established by using the horseradish peroxidase (HRP)/H2O2/halide system in vitro. Neutrophils and lymphocytes, exposed to this system, manifested markedly impaired chemotactic responsiveness and mitogen-induced transformation, respectively. However inclusion of ascorbate with the peroxidative system protected the neutrophils and lymphocytes from these inhibitory effects. Further studies in 3 patients with chronic granulomatous disease (CGD) and 10 patients with bronchial asthma suggested that ascorbate may be of value to improve the primary immunological abnormalities (neutrophil motility and antimicrobial activity) in CGD and the secondary abnormalities (neutrophil motility and lymphocyte transformation) found in some individuals with bronchial asthma.

Uptake of 3,4-dihydroxy[3H]phenylalanine by Mycobacterium leprae isolated from frozen (-80 degrees C) armadillo tissue.

Mycobacterium leprae separated from armadillo tissues stored at -80 degrees C is similar to that from human sources in its ability to take up 3H-labelled 3,4-dihydroxyphenylalanine (DOPA). Several inhibitors were studied which showed complete or partial inhibition of [3H]DOPA uptake. These findings suggest that M. leprae isolated from frozen tissue possesses an active uptake system for [3H]DOPA.

Uptake of 3, 4-dihydroxy(3H)phenylalanine by Mycobacterium leprae isolated from frozen (-80 degrees C) armadillo tissue

Mycobacterium leprae separated from armadillo tissues stored at -80 degrees C is similar to that from human sources in its ability to take up 3H-labelled 3,4-dihydroxyphenylalanine (DOPA). Several inhibitors were studied which showed complete or partial inhibition of [3H]DOPA uptake. These findings suggest that M. leprae isolated from frozen tissue possesses an active uptake system for [3H]DOPA.