Improving cryo-EM grids for amyloid fibrils using interface-active solutions and spectator proteins.

Preparation of cryoelectron microscopy (cryo-EM) grids for imaging of amyloid fibrils is notoriously challenging. The human islet amyloid polypeptide (hIAPP) serves as a notable example, as the majority of reported structures have relied on the use of nonphysiological pH buffers, N-terminal tags, and seeding. This highlights the need for more efficient, reproducible methodologies that can elucidate amyloid fibril structures formed under diverse conditions. In this work, we demonstrate that the distribution of fibrils on cryo-EM grids is predominantly determined by the solution composition, which is critical for the stability of thin vitreous ice films. We discover that, among physiological pH buffers, HEPES uniquely enhances the distribution of fibrils on cryo-EM grids and improves the stability of ice layers. This improvement is attributed to direct interactions between HEPES molecules and hIAPP, effectively minimizing the tendency of hIAPP to form dense clusters in solutions and preventing ice nucleation. Furthermore, we provide additional support for the idea that denatured protein monolayers forming at the interface are also capable of eliciting a surfactant-like effect, leading to improved particle coverage. This phenomenon is illustrated by the addition of nonamyloidogenic rat IAPP (rIAPP) to a solution of preaggregated hIAPP just before the freezing process. The resultant grids, supplemented with this "spectator protein", exhibit notably enhanced coverage and improved ice quality. Unlike conventional surfactants, rIAPP is additionally capable of disentangling the dense clusters formed by hIAPP. By applying the proposed strategies, we have resolved the structure of the dominant hIAPP polymorph, formed in vitro at pH 7.4, to a final resolution of 4 Å. The advances in grid preparation presented in this work hold significant promise for enabling structural determination of amyloid proteins which are particularly resistant to conventional grid preparation techniques.

Co-Aggregation of S100A9 with DOPA and Cyclen-Based Compounds Manifested in Amyloid Fibril Thickening without Altering Rates of Self-Assembly.

The amyloid cascade is central for the neurodegeneration disease pathology, including Alzheimer's and Parkinson's, and remains the focus of much current research. S100A9 protein drives the amyloid-neuroinflammatory cascade in these diseases. DOPA and cyclen-based compounds were used as amyloid modifiers and inhibitors previously, and DOPA is also used as a precursor of dopamine in Parkinson's treatment. Here, by using fluorescence titration experiments we showed that five selected ligands: DOPA-D-H-DOPA, DOPA-H-H-DOPA, DOPA-D-H, DOPA-cyclen, and H-E-cyclen, bind to S100A9 with apparent Kd in the sub-micromolar range. Ligand docking and molecular dynamic simulation showed that all compounds bind to S100A9 in more than one binding site and with different ligand mobility and H-bonds involved in each site, which all together is consistent with the apparent binding determined in fluorescence experiments. By using amyloid kinetic analysis, monitored by thioflavin-T fluorescence, and AFM imaging, we found that S100A9 co-aggregation with these compounds does not hinder amyloid formation but leads to morphological changes in the amyloid fibrils, manifested in fibril thickening. Thicker fibrils were not observed upon fibrillation of S100A9 alone and may influence the amyloid tissue propagation and modulate S100A9 amyloid assembly as part of the amyloid-neuroinflammatory cascade in neurodegenerative diseases.

A comparative assessment of static muscular strength among female operative's working in different handicraft occupations in India.

An ergonomic assessment of static muscular strength like hand grip-and pinch strength in handicraft occupation was made. In total, 170 female participants were selected for the case-control study, out of which 120 handicraft and 50 office workers were examined as exposed and control groups. Their maximum static handgrip- and pinch strength were measured using hand and pinch dynamometers. We analyze our findings to indicate static muscular strength varies significantly due to repetitive use of hand tools. The decrement in pinch grip strength was evident due to long cycle repetitive pinching movements of distal phalanx during hand knotting and pearl drilling. Based on our results we propose the need for ergonomically designed hand tool interventions that may reduce the accumulation of loss in static muscle strength.

Direct Visualization of Model Membrane Remodeling by α-Synuclein Fibrillization.

The interaction of α-synuclein (αS) with membranes is thought to be critical in the etiology of Parkinson's disease. Besides oligomeric αS aggregates that possibly form membrane pores, the aggregation of αS into amyloid fibrils has been reported to disrupt membranes. The mechanism by which aggregation affects the integrity of membranes is, however, unknown. Here, we show that whereas mature αS fibrils only weakly adhere to POPC/POPG giant unilamellar vesicles (GUVs), fibrillization of αS on the membrane results in large-scale membrane remodeling. Fibrils that grow on the vesicle surface stiffen the membrane and make the initially spherical membrane become polyhedral. Additionally, membrane-attached fibrils extract lipids. The lipid extraction and membrane remodeling of growing fibrils can consume the complete bilayer surface and results in loss of vesicle content. These observations suggest that there are several mechanisms by which growing fibrils can disrupt membrane function.

α-Synuclein Oligomers Stabilize Pre-Existing Defects in Supported Bilayers and Propagate Membrane Damage in a Fractal-Like Pattern.

Phospholipid vesicles are commonly used to get insights into the mechanism by which oligomers of amyloidogenic proteins damage membranes. Oligomers of the protein α-synuclein (αS) are thought to create pores in phospholipid vesicles containing a high amount of anionic phospholipids but fail to damage vesicle membranes at low surface charge densities. The current understanding of how αS oligomers damage the membranes is thus incomplete. This incomplete understanding may, in part, result from the choice of model membrane systems. The use of free-standing membranes such as vesicles may interfere with the unraveling of some damage mechanisms because the line tension at the edge of a membrane defect or pore ensures defect closure. Here, we have used supported lipid bilayers (SLBs) of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine (POPC/POPS) to study the membrane damage caused by αS oligomers. Although αS oligomers were not able to initiate the disruption of POPC/POPS vesicles or intact SLBs, oligomers did stabilize and enlarge pre-existing SLB defects. The increased exposure of lipid acyl chains at the edges of defects very likely facilitates membrane-oligomer interactions, resulting in the growth of fractal domains devoid of lipids. Concomitant with the appearance of the fractal membrane damage patterns, lipids appear in solution, directly implicating αS oligomers in the observed lipid extraction. The growth of the membrane damage patterns is not limited by the binding of lipids to the oligomer. The analysis of the shape and growth of the lipid-free domains suggests the involvement of an oligomer-dependent diffusion-limited extraction mechanism. The observed αS oligomer-induced propagation of membrane defects offers new insights into the mechanisms by which αS oligomers can contribute to the loss in membrane integrity.

Optimum iron-pyrophosphate electronic coupling to improve electrochemical water splitting and charge storage.

Tuning the electronic properties of transition metals using pyrophosphate (P2O7) ligand moieties can be a promising approach to improving the electrochemical performance of water electrolyzers and supercapacitors, although such a material's configuration is rarely exposed. Herein, we grow NiP2O7, CoP2O7, and FeP2O7 nanoparticles on conductive Ni-foam using a hydrothermal procedure. The results indicated that, among all the prepared samples, FeP2O7 exhibited outstanding oxygen evolution reaction and hydrogen evolution reaction with the least overpotential of 220 and 241 mV to draw a current density of 10 mA/cm2. Theoretical studies indicate that the optimal electronic coupling of the Fe site with pyrophosphate enhances the overall electronic properties of FeP2O7, thereby enhancing its electrochemical performance in water splitting. Further investigation of these materials found that NiP2O7 had the highest specific capacitance and remarkable cycle stability due to its high crystallinity as compared to FeP2O7, having a higher percentage composition of Ni on the Ni-foam, which allows more Ni to convert into its oxidation states and come back to its original oxidation state during supercapacitor testing. This work shows how to use pyrophosphate moieties to fabricate non-noble metal-based electrode materials to achieve good performance in electrocatalytic splitting water and supercapacitors.

Measuring static muscular strength among female operatives: a cross-sectional comparison in different handicraft occupations.

Purpose. Loss of static muscular strength is the most common work-related problem among handicraft workers involved in hand-intensive jobs. A cross-sectional comparative assessment was carried out to determine the muscular strength among workers involved in the manufacturing of three different crafts: weaving, hand block printing and imitation jewelry. Methods. 120 female operatives were selected, and digital grip dynamometers were used to measure their maximum hand grip and pinch strength. Results. Static muscular strength varies significantly among the different occupational groups of workers. The difference in grip strength in the right and left hands shows that exposure to hand tools for a prolonged period plays a vital role in muscle strength. These findings indicate that static muscular strength varies significantly due to repetitive use of hand tools. The observed values of muscle strength in the dominant hand were significantly lower in jewelry workers while block printing workers had the highest strength. Conclusion. The decrement in pinch grip strength was evident due to long-cycle repetitive pinching movements of the distal phalanx during hand knotting and pearl drilling. This study proposes the need for ergonomically designed hand tool interventions that may reduce the accumulation of loss in static muscle strength.

Natural Compound from Olive Oil Inhibits S100A9 Amyloid Formation and Cytotoxicity: Implications for Preventing Alzheimer's Disease.

Polyphenolic compounds in the Mediterranean diet have received increasing attention due to their protective properties in amyloid neurodegenerative and many other diseases. Here, we have demonstrated for the first time that polyphenol oleuropein aglycone (OleA), which is the most abundant compound in olive oil, has multiple potencies for the inhibition of amyloid self-assembly of pro-inflammatory protein S100A9 and the mitigation of the damaging effect of its amyloids on neuroblastoma SH-SY5Y cells. OleA directly interacts with both native and fibrillar S100A9 as shown by intrinsic fluorescence and molecular dynamic simulation. OleA prevents S100A9 amyloid oligomerization as shown using amyloid oligomer-specific antibodies and cross-ß-sheet formation detected by circular dichroism. It decreases the length of amyloid fibrils measured by atomic force microscopy (AFM) as well as reduces the effective rate of amyloid growth and the overall amyloid load as derived from the kinetic analysis of amyloid formation. OleA disintegrates already preformed fibrils of S100A9, converting them into nonfibrillar and nontoxic aggregates as revealed by amyloid thioflavin-T dye binding, AFM, and cytotoxicity assays. At the cellular level, OleA targets S100A9 amyloids already at the membranes as shown by immunofluorescence and fluorescence resonance energy transfer, significantly reducing the amyloid accumulation in GM1 ganglioside containing membrane rafts. OleA increases overall cell viability when neuroblastoma cells are subjected to the amyloid load and alleviates amyloid-induced intracellular rise of reactive oxidative species and free Ca2+. Since S100A9 is both a pro-inflammatory and amyloidogenic protein, OleA may effectively mitigate the pathological consequences of the S100A9-dependent amyloid-neuroinflammatory cascade as well as provide protection from neurodegeneration, if used within the Mediterranean diet as a potential preventive measure.

Polyoxometalates as Effective Nano-inhibitors of Amyloid Aggregation of Pro-inflammatory S100A9 Protein Involved in Neurodegenerative Diseases.

Pro-inflammatory and amyloidogenic S100A9 protein is central to the amyloid-neuroinflammatory cascade in neurodegenerative diseases. Polyoxometalates (POMs) constitute a diverse group of nanomaterials, which showed potency in amyloid inhibition. Here, we have demonstrated that two selected nanosized niobium POMs, Nb10 and TiNb9, can act as potent inhibitors of S100A9 amyloid assembly. Kinetics analysis based on ThT fluorescence experiments showed that addition of either Nb10 or TiNb9 reduces the S100A9 amyloid formation rate and amyloid quantity. Atomic force microscopy imaging demonstrated the complete absence of long S100A9 amyloid fibrils at increasing concentrations of either POM and the presence of only round-shaped and slightly elongated aggregates. Molecular dynamics simulation revealed that both Nb10 and TiNb9 bind to native S100A9 homo-dimer by forming ionic interactions with the positively charged Lys residue-rich patches on the protein surface. The acrylamide quenching of intrinsic fluorescence showed that POM binding does not perturb the Trp 88 environment. The far and near UV circular dichroism revealed no large-scale perturbation of S100A9 secondary and tertiary structures upon POM binding. These indicate that POM binding involves only local conformational changes in the binding sites. By using intrinsic and 8-anilino-1-naphthalene sulfonate fluorescence titration experiments, we found that POMs bind to S100A9 with a Kd of ca. 2.5 µM. We suggest that the region, including Lys 50 to Lys 54 and characterized by high amyloid propensity, could be the key sequences involved in S1009 amyloid self-assembly. The inhibition and complete hindering of S100A9 amyloid pathways may be used in the therapeutic applications targeting the amyloid-neuroinflammatory cascade in neurodegenerative diseases.

Dissecting the Structural Organization of Multiprotein Amyloid Aggregates Using a Bottom-Up Approach.

Deposition of fibrillar amyloid ß (Aß) in senile plaques is a pathological signature of Alzheimer's disease. However, senile plaques also contain many other components, including a range of different proteins. Although the composition of the plaques can be analyzed in post-mortem tissue, knowledge of the molecular details of these multiprotein inclusions and their assembly processes is limited, which impedes the progress in deciphering the biochemical mechanisms associated with Aß pathology. We describe here a bottom-up approach to monitor how proteins from human cerebrospinal fluid associate with Aß amyloid fibrils to form plaque particles. The method combines flow cytometry and mass spectrometry proteomics and allowed us to identify and quantify 128 components of the captured multiprotein aggregates. The results provide insights into the functional characteristics of the sequestered proteins and reveal distinct interactome responses for the two investigated Aß variants, Aß(1-40) and Aß(1-42). Furthermore, the quantitative data is used to build models of the structural organization of the multiprotein aggregates, which suggests that Aß is not the primary binding target for all the proteins; secondary interactions account for the majority of the assembled components. The study elucidates how different proteins are recruited into senile plaques and establishes a new model system for exploring the pathological mechanisms of Alzheimer's disease from a molecular perspective.

Shape synthesis of an assistive knee exoskeleton device to support knee joint and rehabilitate gait.

Purpose: The assistive knee exoskeleton device is used for supporting the surrounding ligaments, tendons, and muscles of the injured knee joint. Various knee exoskeletons have been discussed; however, their shape synthesis is not reported. This study aims to present the shape synthesis of the assistive knee device. Moreover, four-bar linkage is used for the knee exoskeleton, in this study. Methods: Clinical biomechanical data are adapted from gait database for one gait cycle. Using the clinical gait data, position and static force analyses are performed to obtain a set of orientations and unknown forces. Simultaneously, CAD models are prepared, and the obtained forces are applied to the CAD models of the four-bar linkage knee exoskeleton. Consequently, the threshold is obtained for each component of the knee exoskeleton and the unwanted material below the threshold is removed. Results: A reduction of 45% in the peak actuating force is observed in comparison with the literature. Besides, a total reduction of 21% in the mass of four-bar knee exoskeleton is observed in contrast to the base models when shape synthesis is performed. Conclusions: An assistive knee exoskeleton is developed using the shape synthesis methodology in which four-bar linkage is used. New shapes of thigh and shank attachments are obtained. The developed knee exoskeleton can be used by persons with the injured knee for supporting the ligaments, tendons, and muscles. Besides, control technology can be implemented to make it useful for persons with monoplegia. Implications for rehabilitation Assistive knee exoskeleton devices proved to be an important tool for providing support to injured knee joints. Typically single axis joints are observed in the lower limb exoskeletons which can be replaced with linkage mechanisms to obtain the desired range of motion. In this study, four-bar linkages are used for the knee exoskeleton in which cranks and rockers are connected to the lateral and medial sides of the knee joint, for connecting shank and thigh attachments. Shape synthesis is performed on the components of four-bar exoskeleton through the evaluated reaction forces. The components are assembled to form an assistive knee exoskeleton which can be used by any person with injured knee joint.

Synergistic effects of metal-induced aggregation of human serum albumin.

Exposure to cobalt (Co), chromium (Cr), and nickel (Ni) occurs often via skin contact and from different dental and orthopedic implants. The metal ions bind to proteins, which may induce structural changes and aggregation, with different medical consequences. We investigated human serum albumin (HSA) aggregation in the presence of CoII, CrIII, and/or NiII ions and/or their nanoparticle precipitates by using scattering, spectroscopic, and imaging techniques, at simulated physiological conditions (phosphate buffered saline - PBS, pH 7.3) using metal salts that did not affect the pH, and at HSA:metal molar ratios of up to 1:8. Co ions formed some solid nanoparticles in PBS at the investigated conditions, as determined by nanoparticle tracking analysis, but the CrIII anions and NiII ions remained fully soluble. It was found that all metal ions induced HSA aggregation, and this effect was significantly enhanced when a mixture of all three metal ions was present instead of any single type of ion. Thus, the metal ions induce aggregation synergistically. HSA aggregates formed linear structures on a mica surface in the presence of CrIII ions. A clear tendency of aggregation and linearly aligned aggregates was seen in the presence of all three metal ions. Spectroscopic investigations indicated that the majority of the HSA molecules maintained their alpha helical secondary structure and conformation. This study highlights the importance of synergistic effects of metal ions and/or their precipitates on protein aggregation, which are highly relevant for implant materials and common exposures to metals.

Intrinsically disordered protein as carbon nanotube dispersant: How dynamic interactions lead to excellent colloidal stability.

The rich pool of protein conformations combined with the dimensions and properties of carbon nanotubes create new possibilities in functional materials and nanomedicine. Here, the intrinsically disordered protein α-synuclein is explored as a dispersant of single-walled carbon nanotubes (SWNTs) in water. We use a range of spectroscopic methods to quantify the amount of dispersed SWNT and to elucidate the binding mode of α-synuclein to SWNT. The dispersion ability of α-synuclein is good even with mild sonication and the obtained dispersion is very stable over time. The whole polypeptide chain is involved in the interaction accompanied by a fraction of the chain changing into a helical structure upon binding. Similar to other dispersants, we observe that only a small fraction (15-20%) of α-synuclein is adsorbed on the SWNT surface with an average residence time below 10 ms.

A novel gait-based synthesis procedure for the design of 4-bar exoskeleton with natural trajectories.

BACKGROUND/OBJECTIVE: Human walking involves the coordination of brain, nerves, and muscles. A disturbance in their coordination may result in gait disorder. The gait disorder may be treated through manually assisted gait training or with the aid of assistive devices/robotic devices. These robotic devices involve mechanisms which are synthesized using complex conventional procedures. Therefore, in this study, a new gait-based synthesis procedure is proposed, which simplifies the mechanism synthesis and helps to develop a mechanism which can be used in rehabilitation devices, bipeds, etc. METHODS: This article presents a novel procedure for the synthesis of 4-bar linkage using the natural gait trajectories. As opposed to the conventional synthesis procedures, in this procedure, a global reference frame is considered, which allows the use of hip trajectory while moving. Moreover, this method is divided into two stages, and five precision points are considered on the hip trajectory in each stage. In the first stage, the 4-bar linkage is designed, thereafter, the configurations of the linkage for the remaining precision points are determined in the second stage. The proposed synthesis procedure reduces the complexity involved in the synthesis and helps in the simplification of the problem formulation. A two-stage optimization problem is formulated for minimizing the error between the generated and desired hip trajectories. Two nature-inspired algorithms are used for solving the optimization problem. The obtained best results are presented, and the designed linkage is simulated in MATLAB. RESULTS: The best design of the linkage is obtained using particle swarm optimization. The trajectories generated by the designed linkage using the proposed methodology can accurately track the desired path, which indicates that designed linkage can achieve all the orientations required during walking. The positions of a whole lower limb at all the desired precision points are demonstrated by stick diagram for one gait. CONCLUSION: The proposed methodology has reduced the complexity of synthesis procedures and used optimization techniques to obtain a feasible design of the mechanism. The stick diagram of the designed mechanism obtained using the proposed method indicates that the designed mechanism can walk smoothly. Hence, the designed mechanism can be used in the rehabilitation devices. Furthermore, a conceptual design of an exoskeleton knee is also presented. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: Many hospitals and individuals have used the immobile and portable rehabilitation devices. These devices involve mechanisms, and the design of mechanism plays a vital role in the functioning of these devices; therefore, we have developed a new synthesis procedure for the design of the mechanism. Besides synthesis procedure, a mechanism is developed that can be used in the rehabilitation devices, bipeds, exoskeletons, etc., to benefit the society.

Tubercular uterocutaneous fistula after caesarean section: A case report.

A 29-year-old patient had undergone an elective lower-segment caesarean section (LSCS) five months previously at a district hospital. The operation and the immediate postoperative period were uneventful. After five months she presented back with a fistulous opening. A fistulogram revealed a connection between the uterus and the skin. Fistulous tract excision was planned. Intraoperatively there was communication between the skin and the uterine cavity, with extensive necrosis of the uterine wall. The patient gave her informed consent for excision of the fistulous tract and/or total abdominal hysterectomy. During surgery, it was deemed that there was no scope for excision, so the decision was made for a total abdominal hysterectomy. Histopathological examination confirmed tuberculosis and the patient responded well to anti-tubercular drugs. This case report describes a rare presentation of tubercular uterocutaneous fistula after caesarean section.

An investigation on effects of amputee's physiological parameters on maximum pressure developed at the prosthetic socket interface using artificial neural network.

Technological advances in prosthetics have attracted the curiosity of researchers in monitoring design and developments of the sockets to sustain maximum pressure without any soft tissue damage, skin breakdown, and painful sores. Numerous studies have been reported in the area of pressure measurement at the limb/socket interface, though, the relation between amputee's physiological parameters and the pressure developed at the limb/socket interface is still not studied. Therefore, the purpose of this work is to investigate the effects of patient-specific physiological parameters viz. height, weight, and stump length on the pressure development at the transtibial prosthetic limb/socket interface. Initially, the pressure values at the limb/socket interface were clinically measured during stance and walking conditions for different patients using strain gauges placed at critical locations of the stump. The measured maximum pressure data related to patient's physiological parameters was used to develop an artificial neural network (ANN) model. The effects of physiological parameters on the pressure development at the limb/socket interface were examined using the ANN model. The analyzed results indicated that the weight and stump length significantly affects the maximum pressure values. The outcomes of this work could be an important platform for the design and development of patient-specific prosthetic socket which can endure the maximum pressure conditions at stance and ambulation conditions.

Lipid Head Group Charge and Fatty Acid Configuration Dictate Liposome Mobility in Neurofilament Networks.

Intermediate filaments constitute a class of biopolymers whose function is still poorly understood. One example for such intermediate filaments is given by neurofilaments, large macromolecules that fill the axon of neurons. Here, reconstituted networks of purified porcine neurofilaments are studied and the diffusion behavior of different nanoparticles in the biopolymer network is evaluated. A strong dependence of particle diffusion on the charge state of the particles, and - for liposomes - also on the fatty acid configuration of lipids is observed. The results suggest that both electrostatic and hydrophobic interactions contribute to nanoparticle trapping in neurofilament networks, and that the latter is enabled by lipids with an inverted cone geometry which grant access to the hydrophobic core of the liposome shell.

Membrane interactions and fibrillization of α-synuclein play an essential role in membrane disruption.

We studied α-synuclein (αS) aggregation in giant vesicles, and observed dramatic membrane disintegration, as well as lipid incorporation into micrometer-sized suprafibrillar aggregates. In the presence of dye-filled vesicles, dye leakage and fibrillization happen concurrently. However, growing fibrils do not impair the integrity of phospholipid vesicles that have a low affinity for αS. Seeding αS aggregation accelerates dye leakage, indicating that oligomeric species are not required to explain the observed effect. The evolving picture suggests that fibrils that appear in solution bind membranes and recruit membrane-bound monomers, resulting in lipid extraction, membrane destabilization and the formation of lipid-containing suprafibrillar aggregates.

Sigmoid vaginoplasty: long-term results.

OBJECTIVES: To evaluate the long-term results of sigmoid vaginoplasty for Mayer-Rokitansky-Kuster-Hauser syndrome. The social and psychological acceptance of the procedure is also discussed in terms of a developing country scenario. METHODS: A total of 14 patients with Mayer-Rokitansky-Kuster-Hauser syndrome were treated at our institute from January 1995 to December 2004. Sigmoid vaginoplasty was performed in all patients. The procedure was performed using a combined abdominoperineal approach. Dissection was done between the urethra and rectum to create a bed for the neovaginal colon conduit. A 10-cm segment of sigmoid colon was raised on its vascular pedicle, delivered through the abdominoperineal tunnel, and fixed to the vaginal pit incision. The patient records were reviewed for surgical technique and postoperative complications. Patients underwent a personal interview to assess the postoperative results, social acceptance of the procedure, and sexual satisfaction. RESULTS: The mean patient age at surgery was 16.8 years. The patients who underwent sigmoid vaginoplasty had good cosmetic results without the need for routine dilation or the problem of excessive mucus production. The postoperative morbidity was minimal. During a mean follow-up of 4.1 years, no stenosis or colitis was encountered. The subjective satisfaction rate with the surgical outcomes in all the patients was 8.01 on a scale of 0 to 10 (0, very disappointed to 10, satisfied). CONCLUSIONS: Sigmoid vaginoplasty is an effective treatment for patients with vaginal atresia. Timed vaginal reconstruction in these patients allows for a better quality of life and social acceptance. It also enables the patient to lead a near-normal sexual life, with high satisfaction rates.

Association of vitamin-D and calcitonin receptor gene polymorphism in paediatric nephrolithiasis.

We investigated the role of vitamin-D receptor gene (VDR) and calcitonin receptor (CTR) gene polymorphism in childhood nephrolithiasis in the north Indian population. A control group of 60 healthy paediatric individuals (age range 4-16 years) and 50 paediatric patients (age range 2-14 years) with kidney stones were examined. Polymorphism in both genes (VDR and CTR) was detected by using PCR-based restriction analysis. There was a statistically significant difference between the two groups for the genotypes of the VDR gene Fok-I polymorphism (P =0.007) and the CTR gene (P =0.048) polymorphism. The odds ratio (95% Confidence Interval) for the C allele in those at risk of stone disease was 1.83 (0.82-4.09) in VDR gene polymorphism and 1.99 (0.90-4.39) in the case of CTR gene polymorphism. Our results suggest that the effects of VDR (Fok-I) and CTR gene polymorphism contribute to the understanding of the pathogenesis of urinary calculi. It is also suggestive of a potential candidate gene in the search for genetic causes of paediatric calcium oxalate nephrolithiasis.