Probing High-order Protein Complexes Using Native Mass Spectrometry and Hydrogen/Deuterium Exchange Mass Spectrometry: A Case Study Using Fresh and Commercial Hemoglobin Samples.

Native mass spectrometry (MS) analysis of protein complexes is highly susceptible to matrix effect, and addressing this predicament using buffer exchange is a common approach. Nevertheless, optimization of the buffer exchange protocol is not trivial. With the use of hemoglobin (Hb) as the model entity, it was discovered that the native mass spectrum of protein assembly is highly dependent on the buffer-exchange protocol. Given the dependence of native MS on the purification protocol, this work attempts to use hydrogen/deuterium exchange mass spectrometry (HDX-MS) for comparative studies of hemoglobin complexes in untreated fresh and commercial samples. The information obtained from the HDX study was found to correlate well with the native mass spectrometry analysis of the properly buffer-exchanged Hb samples. Both native MS and HDX-MS showed that the fresh Hb sample has retained the expected tetrameric structure, whereas the commercial Hb has largely been denatured to the dimeric form. These findings prove the complementarity of native MS and HDX-MS in the analysis of high-order protein complexes and stress the necessity to validate the integrity of the high-order structures of the proteins prior to the use of the protein samples for other biomedical studies.

A millimeter water-in-oil droplet as an alternative back exchange prevention strategy for hydrogen/deuterium exchange mass spectrometry of peptides/proteins.

Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a versatile bioanalytical technique for protein analysis. Since the reliability of HDX-MS analysis considerably depends on the retention of deuterium labels in the post-labeling workflow, deuterium/hydrogen (D/H) back exchange prevention strategies, including decreasing the pH, temperature, and exposure time to protic sources of the deuterated samples, are widely adopted in the conventional HDX-MS protocol. Herein, an alternative and effective back exchange prevention strategy based on the encapsulation of a millimeter droplet of a labeled peptide solution in a water-immiscible organic solvent (cyclohexane) is proposed. Cyclohexane was used to prevent the undesirable uptake of water by the droplet from the atmospheric vapor through the air-water interface. Using the pepsin digest of deuterated myoglobin, our results show that back exchange kinetics of deuterated peptides is retarded in a millimeter droplet as compared to that in the bulk solution. Performing pepsin digestion directly in a water-in-oil droplet at room temperature (18-21 °C) was found to preserve more deuterium labels than that in the bulk digestion with an ice-water bath. Based on the present findings, it is proposed that keeping deuterated peptides in the form of water-in-oil droplets during the post-labelling workflow will facilitate the preservation of deuterium labels on the peptide backbone and thereby enhance the reliability of the H/D exchange data.

Peptide Oxidation Induced by Liquid-Solid Contact Electrification as Revealed in Liquid Microjunction-Surface Sampling Probe Mass Spectrometry.

Reproducible peptide oxidation was observed using a homebuilt liquid microjunction-surface sampling probe (LMJ-SSP) platform for analyzing peptide standards. Although electrochemical oxidation and corona discharges have previously been associated with analyte oxidation in electrospray ionization (ESI) and ESI-related ambient ionization mass spectrometry (MS) methods, they were unlikely the causes for the peptide oxidation observed in the LMJ-SSP studies. A systematic investigation demonstrated that analyte oxidation was induced during the droplet drying on a solid surface through liquid-solid electrification processes. To minimize unwanted analyte oxidation, the water content in the sample solution should be decreased and the use of hydroxyl-functionalized substrates, such as glass slides, should be avoided. In addition, if water is an essential solvent component, adding an antioxidant, such as ascorbic acid, to the sample solution before droplet evaporation on the solid surface could lower the percentage of analyte oxidation. The present findings apply to all the MS methods that involve drying microliters of sample solution onto a suitable substrate in their sample preparation protocols.

CaptiveSpray Differential Ion Mobility Spectrometry Device with Enhanced Ion Transmission and Improved Resolving Power.

A performance enhanced CaptiveSpray differential ion mobility device was designed and constructed by incorporating a circular channel and a gas flow homogenizing channel (GFHC) between the CaptiveSpray ion source and planar differential ion mobility spectrometry (DMS). The GFHC was used to reduce gas flow heterogeneity prior to the entrance of the DMS device. The optimal flared entrance greatly reduces gas flow velocity at the inlet region owing to its relatively large gas inlet interface, which assists in reducing disparities between the minimum and maximum gas velocity along the x-axis. The circular electrode was machined with channels along the x- and y-axis for the passage of auxiliary gas and was applied with a potential to focus the incoming ions from the CaptiveSpray source into the DMS channel. Using reserpine as a reference standard, substantial signal enhancement was achieved with a concomitant reduction of the peak width in the ionogram.

Tandem Mass Spectrometry for Structural Characterization of Doubly-Charged N-Linked Glycopeptides.

Three dissociation methods, including collision-induced dissociation (CID), electron capture dissociation (ECD), and electronic excitation dissociation (EED), were systematically compared for structural characterization of doubly charged glycopeptide. CID produced distinctively different tandem mass spectra for glycopeptide adducted with different charge carriers. Protonated species produced mainly glycosidic cleavages in high abundance. CID of magnesiated glycopeptide formed more cross-ring cleavages, whereas doubly sodiated species produced cleavages at both glycan and peptide moieties. The effect of charge carriers on the fragmentation in ECD and EED was lower than that in CID. ECD produced mainly peptide backbone cleavages but limited cleavages at the glycan moiety, whereas EED of glycopeptide resulted in extensive fragmentation throughout the molecular ion regardless of the charge carriers. Magnesiated species gave, however, more cross-ring cleavages than other charge carriers did. These results demonstrated that EED of magnesiated species could be used as a one-step dissociation method for comprehensive structural analysis of glycopeptides.

Ideal implant choice for proximal interphalangeal joint arthrodesis in hammer toe/claw toe deformity correction: A systematic review.

Intramedullary devices have been developed to reduce the problems associated with Kirschner (K)-wire fixation in proximal interphalangeal joint (PIPJ) arthrodesis. The purpose of this systematic review is to compare the surgical outcomes of K-wires versus novel internal fixation devices in PIPJ arthrodesis in claw/hammer toe surgery. The databases searched were PubMed, Scopus, Cochrane, and Embase with keywords "claw toe OR hammer toe" AND "proximal interphalangeal OR PIP" AND "fusion OR arthrodesis." Clinical trials published in English with evidence levels I, II, and III were included. Five studies, including one randomized controlled trial and four case-controlled studies, were identified to meet the inclusion criteria. Overall, the studies showed promising results in union rates using the novel internal devices compared to K-wires. However, the novel internal devices seem not to present advantages in clinical parameters such as pain levels, patient satisfaction, foot-related function, or surgical complication rates.

Endoscopic Achilles tendon shortening.

Healing in continuity with lengthening of the Achilles tendon is a disabling complication after Achilles tendon rupture. This results in weakness of ankle plantarflexion and a non-propulsive gait on the affected side. The elongated tendon may have to be shortened or reconstructed to re-establish the length and the physiologic tension in triceps surae muscle and allow restoration of push off. This technical note describes the details of endoscopic shortening of the Achilles tendon.

Reliability and validity analysis of the open-source Chinese Foot and Ankle Outcome Score (FAOS).

AIM: Develop the first reliable and validated open-source outcome scoring system in the Chinese language for foot and ankle problems. METHODS: Translation of the English FAOS into Chinese following regular protocols. First, two forward-translations were created separately, these were then combined into a preliminary version by an expert committee, and was subsequently back-translated into English. The process was repeated until the original and back translations were congruent. This version was then field tested on actual patients who provided feedback for modification. The final Chinese FAOS version was then tested for reliability and validity. Reliability analysis was performed on 20 subjects while validity analysis was performed on 50 subjects. Tools used to validate the Chinese FAOS were the SF36 and Pain Numeric Rating Scale (NRS). Internal consistency between the FAOS subgroups was measured using Cronbach's alpha. Spearman's correlation was calculated between each subgroup in the FAOS, SF36 and NRS. RESULTS: The Chinese FAOS passed both reliability and validity testing; meaning it is reliable, internally consistent and correlates positively with the SF36 and the NRS. DISCUSSION AND CONCLUSIONS: The Chinese FAOS is a free, open-source scoring system that can be used to provide a relatively standardised outcome measure for foot and ankle studies.

Arthroscopic approach to the spring (calcaneonavicular) ligament.

BACKGROUND: This research studied the safety and efficacy of a new portal to the spring ligament. This portal is located just plantar to the insertion of the posterior tibial tendon and above the fibrous septum between the posterior tibial and the flexor digitorum longus tendons. METHODS: Twelve fresh frozen foot and ankle specimens were used. The distance between the accessory medial portal and the medial plantar nerve was measured. The relation between the medial plantar nerve and the spring ligament was studied. The depth that can be reached through the portal was also assessed. RESULTS: The average distance between the insertion point of the 3mm diameter metal rod and the medial plantar nerve was 20(6-27)mm. The medial plantar nerve located at lateral third of the ligament in 8 specimens (67%), middle third in 2 specimens (17%) and medial third in 2 specimens (17%). The tip of rod can reach Zone A in all specimens. CONCLUSION: This study demonstrated that arthroscopic approach and repair of the spring ligament can injure the medial plantar nerve. CLINICAL RELEVANCE: The clinical relevance of this cadaver study is that it confirmed the feasibility of arthroscopic approach to the whole span of the spring ligament and alerted the potential risk of injury to the medial plantar nerve during arthroscopic assisted repair of the ligament.

Avulsion fracture of the posterosuperior tuberosity of the calcaneus managed with lag screw fixation.

BACKGROUND: To review the efficacy of lag screw fixation in management of avulsion fracture of the posterosuperior tuberosity of the calcaneus. METHODS: Since 2002, thirteen patients with displaced fracture of the posterior tuberosity of the calcaneus were treated with emergency reduction of the fracture and lag screw fixation. The medical records and radiographs of the patients were reviewed and the patients were assessed according to the Kerr calcaneal fracture scoring system during the latest follow up. RESULTS: There was no skin necrosis, but one wound dehisced in a patient with unstable diabetes and hypothyroidism. All fractures healed, but two had separation of the fracture fragments after the plaster was removed, both of them were elderly osteoporotic patients. The overall average calcaneal score was 93 (range, 77-100). The average calcaneal score of the patients with closed reduction was 91 (range, 77-100). The average calcaneal score of the patients with open reduction was 94 (range, 79-100). Complications occurred in seven patients (54%). CONCLUSIONS: Percutaneous or open reduction of the avulsion fracture of the posterosuperior tuberosity of the calcaneus together with lag screw fixation and equinus short leg cast immobilization can provide good results without skin necrosis. The surgeon should pay attention of the details of the operation and the rehabilitation program in order to minimize the complications.

5-year results of the 1.5cm incision Achilles tendon repair.

BACKGROUND: To study a hypothesis that the cost-effective 1.5cm medial incision Achilles tendon repair technique will provide good functional outcomes which are maintained for over 5 years. METHOD: Prospective study of 12 consecutive cases with a minimal 5-year follow-up were recruited from April 2008 to November 2010. Cases whom were mentally incompetent or those which required concomitant procedures were excluded. Outcomes measures included the numeric pain rating scale, motor power strength, range of motion, functional scoring using the AOFAS hindfoot score and patient's self-assessment using the Foot and Ankle Outcome Score (FAOS). RESULT: No re-ruptures or sural nerve injured were identified after a minimal 5-year follow-up. Pain was minimal at 0.5/10, calf power was 5/5 and ankle range was good (plantarflexion: 38°/dorsiflexion: 21°). The AOFAS hindfoot score was 97.4 and all 5 sub-categories of the Foot and Ankle Outcome Score (FAOS) were good. CONCLUSION: The 1.5cm medial incision repair of the Achilles tendon is an economically sound surgical technique, with minimal complications, which gives good medium length functional outcomes. LEVEL OF EVIDENCE: IV.

A large solitary fibroma of the deep compartment of the sole.

A 50-year-old lady presented with a right foot mass and pain for 1year. Magnetic resonance imaging showed a 3.9×3.2×5cm mass on plantar side of the forefoot deep to the major flexor tendons and plantar aponeurosis and inferior to 1st-4th metatarsals. The mass extended dorsally through the intermetatarsal space to the foot dorsum. Ultrasound guided biopsy was performed, which confirmed it was a fibroma. This case was further complicated by pathological fracture of the 3rd metatarsal. It was resected through a dorsal incision and surgical dislocation of the 2nd and 3rd tarsometatarsal joints. Magnetic resonance imaging was repeated 1year later and showed complete resection of the lesion without recurrence.

Modified double plantar plate tenodesis.

BACKGROUND: Metatarsophalangeal joint instability of the lesser toe can cause acquired toe deformity. Plantar plate deficiency is the major pathology. Plantar plate repair can stabilize the joint but may result in iatrogenic transverse plane toe deformity in correction of claw toe deformity. Limited toe extension can be resulted after correction of crossover toe deformity by plantar plate tenodesis and extensor digitorum brevis transfer. A modification of the technique is proposed. MATERIALS AND METHODS: The clinical outcomes of 10 patients with the modified procedure performed were assessed. RESULTS: The correction was full in all toes with no recurrence. CONCLUSION: The modified technique can stabilize the metatarsophalangeal joint and correct lesser toe deformity without the need of tendon transfer, osteotomy or sophisticated instrumentation.

Posterior ankle and hindfoot endoscopy: A cadaveric study.

BACKGROUND: The list of indications of posterior ankle endoscopy is expanding and includes various soft tissue and bony pathologies of the posterior ankle. Some of the indications, e.g. release of frozen ankle, debridement of posteromedial soft tissue impingement of the ankle and debridement or fixation of the posteromedial osteochondral lesion of the talus, require approach to the posterior ankle medial to the flexor hallucis longus tendon. The purpose of this study was to assess the risk of injury to the posterior tibial neurovascular bundle during posterior ankle endoscopy. METHODS: Fourteen fresh frozen foot and ankle specimens were used. A metal rod was inserted into the posteromedial, posterolateral and modified posteromedial portals and touched the medial border of the posterolateral talar tubercle and the posteromedial corner of the ankle mortise in turn. The neurovascular bundle and FHL tendon were examined for any kink. RESULTS: The neurovascular bundle was kinked in all specimens (100%) with the rod at the posteromedial corner of the ankle mortise through the posteromedial portal and was kinked in 11 specimens (79%) with the rod through the modified posteromedial portal. The neurovascular bundle was kinked in 1 specimen (7%) with the rod through the posterolateral portal. CONCLUSIONS: The neurovascular bundle was at risk during instrumentation of the posteromedial ankle through the posteromedial portal but was safe through the posterolateral portal.

Dorsalis pedis psuedoaneurysm: A complication followed extensor tendoscopy of the ankle in a non-tuberculosis patient with tenosynovitis with rice body formation.

Tenosynovitis with rice body formation is very rare in the ankle region. A case of this condition in a non-tuberculosis patient was presented. He was treated by extensor tendoscopy of the ankle. The tenosynovitis subsided after the procedure. However, it was complicated by formation of a dorsalis pedis pseudoaneurysm.

Subperiosteal Hematoma of the Ankle.

INTRODUCTION: Periosteal reaction has a long list of differential diagnoses ranging from trauma, infection, metabolic disease to malignancy. The morphology of periosteal reaction shown in imaging studies helps to narrow down the list of differential diagnoses. CASE REPORT: A 25 year old gentleman had an inversion injury to his left ankle. He complained of lateral ankle and posterior heel pain and swelling after the injury. Radiograph of his left ankle revealed solid, smooth periosteal reaction at posterior aspect of left distal tibia. MRI showed periosteal reaction at the corresponding site, which was better demonstrated in CT scan. Follow up MRI and CT showed maturation of the new bone formation at the site of periosteal reaction. Findings were compatible with subperiosteal hematoma formation from injury, which ossified with time. CONCLUSION: Smooth, thick periosteal reaction favours benign process, while interrupted pattern is an alarming feature for more aggressive causes.

Stabilization of medial longitudinal foot arch by peroneus longus transfer.

The mechanical integrity of the medial longitudinal arch depends on the dynamic support of muscles and the static support of ligaments. Although the posterior tibial tendon is the main dynamic stabilizer of the arch, the static structures provide the most support especially while the person is standing. After rupture of the posterior tibial tendon, the spring ligament may be compromised under increased stress and leads to talar derotation and peritalar subluxation. Surgical repair of the spring ligament has become an important adjunct to treating posterior tibial tendon abnormalities. A technique of peroneus longus transfer to augment the static stabilizers of the medial column is described in this article.

The safety of the posterior ankle arthroscopy in management of posterior ankle impingement: A cadaveric study.

BACKGROUND: Arthroscopic management of the posterior ankle impingement with the patient in supine position has the advantage of dealing with anterior ankle pathology at the same time without the need to change position of the patient. This study aims at evaluation of the safety of portal establishment and instrumentation of this technique. METHODS: Sixteen fresh-frozen cadaver specimens were used. The relationships of the posteromedial and posterolateral portals to the adjacent tendons and nerves and the relationship of the coaxial portal tract with the posterior ankle capsule and the flexor hallucis longus tendon were studied. RESULT: Angle θ1 between the intermalleolar line and the posterior ankle coaxial portal tract averaged 1° (-10° to 22°). Angle θ2 between the intermalleolar line and the metal rod where the neurovascular bundle started to move averaged 19° (10° to 30°). Angle θ3 between the intermalleolar line and the metal rod where it reached the lateral border of the Achilles tendon was larger than angle θ2 in all specimens. The angle of safety (θs) averaged 18° (-1° to 26°). CONCLUSIONS: Injury to the tendon, nerves or vessels is possible during establishment of the portals and resection of the os trigonum.

First metatarsophalangeal arthroscopy in patients with post-traumatic hallux valgus.

Post-traumatic hallux valgus is relatively rare and has been reported after rupture of the medial collateral ligament of the first metatarsophalangeal (MTP-1) joint; Lisfranc joint injury; turf toe injury; medial plantar nerve entrapment secondary to tibial fracture or first metatarsal fracture. Post-traumatic hallux valgus after medial collateral ligament injury has a high incidence of MTP-1 pathology. Detailed history and clinical examination can facilitate differentiation of the source(s) of the patient's symptoms and assist accurate formulation of the surgical plan. First, MTP arthroscopy is a feasible diagnostic and therapeutic tool to manage the MTP-1 joint pain in hallux valgus following injury to the MTP-1 joint.

Endoscopic curettage and bone grafting of the enchondroma of the proximal phalanx of the great toe.

Enchondromas are the most commonly found benign tumors of the small bones of the foot, mainly involve the phalanges and metatarsals. They are usually asymptomatic, and are found as an incidental finding on routine X-rays. Surgical intervention is indicated in symptomatic lesions as well as larger lesions (greater than 3-4cm) even if these lesions are asymptomatic. Enchondroma most often can be adequately treated with intralesional curettage and bone grafting. A technique of endoscopic curettage and bone grafting of enchondroma of the proximal phalanx of the hallux is described. This has the advantage of minimally invasive surgery of better cosmesis, less surgical trauma and preservation of the cortical integrity.