Primary gastric squamous cell carcinoma - a case report of diagnosis, treatment, histological findings and follow-up.

Primary squamous cell carcinoma (PSCC) of the stomach is a rare oncologic disease with a reported incidence of only 0.04%. Therefore, there are no evidence-based recommendations for treatment. We present the case of a 73-year-old patient with PSSC. Multivisceral resection had to be performed to achieve tumor-free resection margins. Preoperative investigations, surgical treatment, histologic criteria and findings, and follow-up are presented.

Use of Indocyanine Green Fluorescence Angiography and Prophylactic Endoscopic Stenting in Esophagectomy for Acute Esophageal Necrosis.

Acute esophageal necrosis (AEN) is a rare condition with high mortality. We describe a case of an AEN accompanied with multiple esophageal perforations in a 40-year-old patient. This was addressed via emergency Ivor-Lewis-esophagectomy with intraoperative Indocyanine-Green-Angiography (ICGA) and prophylactic placement of a self-expanding metal-stent (SEMS).

[Functional Results after Laparoscopic versus Robot-assisted Sigmoid Resection in Diverticulitis]. / Funktionelle Ergebnisse nach laparoskopischer vs. roboterassistierter Sigmaresektion bei Divertikulitis.

BACKGROUND: Sigmoid resection in diverticulitis is one of the most frequently performed colonic operations. A minimally invasive approach by laparoscopy is the surgical gold standard. For a few years now, sigmoid resections have also been performed robotically (da Vinci® System). It is unclear whether there are relevant differences between the two procedures in terms of functional outcome. METHODS: A postoperative follow-up was performed on all patients who underwent laparoscopic or robotic sigmoid resection for diverticulitis between November 2013 and November 2018, with a minimum interval of 6 months between surgery. Continence disorders, the development of symptoms compared to preoperative, changes in bowel movement (constipation, diarrhoea), impairment in daily life and pain were recorded. Differences between the groups were checked for statistical significance using the chi-square test. RESULTS: During the study period, a total of 106 minimally invasive sigmoid resections for diverticulitis were performed (laparoscopic: n = 46, robot-assisted: n = 60). Of these, 74 patients (70%; laparoscopic: n = 28, robot-assisted: n = 46) answered the questionnaire and were included in the evaluation. Continence disorders were reported in a total of 22% of cases. There was no statistically significant difference between the groups in any of the variables surveyed. CONCLUSION: With regard to functional results after minimally invasive sigmoid resection for diverticulitis, laparoscopic and robot-assisted sigmoid resection can be considered equivalent procedures.

Tuning in C-nociceptors to reveal mechanisms in chronic neuropathic pain.

OBJECTIVE: Develop and validate a low-intensity sinusoidal electrical stimulation paradigm to preferentially activate C-fibers in human skin. METHODS: Sinusoidal transcutaneous stimulation (4Hz) was assessed psychophysically in healthy volunteers (n = 14) and neuropathic pain patients (n = 9). Pursuing laser Doppler imaging and single nociceptor recordings in vivo in humans (microneurography) and pigs confirmed the activation of "silent" C-nociceptors. Synchronized C-fiber compound action potentials were evoked in isolated human nerve fascicles in vitro. Live cell imaging of L4 dorsal root ganglia in anesthetized mice verified the recruitment of small-diameter neurons during transcutaneous 4-Hz stimulation of the hindpaw (0.4mA). RESULTS: Transcutaneous sinusoidal current (0.05-0.4mA, 4Hz) activated "polymodal" C-fibers (50% at ∼0.03mA) and "silent" nociceptors (50% at ∼0.04mA), intensities substantially lower than that required with transcutaneous 1-ms rectangular pulses ("polymodal" ∼3mA, "silent" ∼50mA). The stimulation induced delayed burning (nonpulsating) pain and a pronounced axon-reflex erythema, both indicative of C-nociceptor activation. Pain ratings to repetitive stimulation (1 minute, 4Hz) adapted in healthy volunteers by Numeric Rating Scale (NRS) -3 and nonpainful skin sites of neuropathic pain patients by NRS -0.5, whereas pain even increased in painful neuropathic skin by approximately NRS +2. INTERPRETATION: Sinusoidal electrical stimulation at 4Hz enables preferential activation of C-nociceptors in pig and human skin that accommodates during ongoing (1-minute) stimulation. Absence of such accommodation in neuropathic pain patients suggest axonal hyperexcitability that could be predictive of alterations in peripheral nociceptor encoding and offer a potential therapeutic entry point for topical analgesic treatment. Ann Neurol 2018;83:945-957.

Extensive Chemical Modifications in the Primary Protein Structure of IgG1 Subvisible Particles Are Necessary for Breaking Immune Tolerance.

A current concern with the use of therapeutic proteins is the likely presence of aggregates and submicrometer, subvisible, and visible particles. It has been proposed that aggregates and particles may lead to unwanted increases in the immune response with a possible impact on safety or efficacy. The aim of this study was thus to evaluate the ability of subvisible particles of a therapeutic antibody to break immune tolerance in an IgG1 transgenic mouse model and to understand the particle attributes that might play a role in this process. We investigated the immunogenic properties of subvisible particles (unfractionated, mixed populations, and well-defined particle size fractions) using a transgenic mouse model expressing a mini-repertoire of human IgG1 (hIgG1 tg). Immunization with proteinaceous subvisible particles generated by artificial stress conditions demonstrated that only subvisible particles bearing very extensive chemical modifications within the primary amino acid structure could break immune tolerance in the hIgG1 transgenic mouse model. Protein particles exhibiting low levels of chemical modification were not immunogenic in this model.

[A rare case of coecal volvulus after colonoskopy due to a mobile coekum - diagnosis, surgical therapy and postoperative complications]. / Ein seltener Fall eines Volvulus des Coecums nach Koloskopie bei Coecum mobile ­ Darstellung von Diagnostik, operativer Therapie und postoperativen Komplikationen.

Colonoscopy, either performed as screening or as a therapeutic proceedure, is, in general, very safe with only a few cases of serious complications. Most cases of bleeding after endoscopic polypectomy can be safely managed endoscopically. The rare cases of colonic perforations still have to be managed by surgical intervention. The postpolypectomy-coagulation syndrome and a cecal volvulus are very rare complicatoions after colonoscopy. In the current manuscript, we describe a rare case of a cecal volvulus after routine colonoscopy due to an unknown mobile coecum as a predisposition. We discuss the endoscopic, clinical and radiological findings of the patient. Moreover, we describe the performed surgical procedure and the further clinical course of the patient. A cecal volvulus should always be considered as a possible rare, but serious, complication in the differential diagnosis of abdominal pain after colonoscopy. The standard therapy of a cecal volvulus is the right hemicolectomy. As an alternative, a coecopexy without resection could be performed as long as the colonic wall is still vital.

[The Relevance of Surgeon-performed Ultrasound For the Detection of Acute Appendicitis: a Review of the Literature and a Practical Diagnostic Algorithm]. / Stellenwert der chirurgischen Sonografie in der Diagnostik der akuten Appendizitis: Literaturübersicht und ein praxisnaher Diagnosealgorithmus.

Background The relevance of ultrasound (US) in diagnosing acute appendicitis is controversial. The validity of US in comparison with other imaging techniques, such as computed tomography (CT) and magnetic resonance imaging (MRI), is unclear, as is the difference between surgeon-performed ultrasound (SPUS) and radiologist-performed ultrasound (RPUS). On the basis of a review of current literature, our study aimed to develop a US-based algorithm to simplify the choice between additional diagnostic measures and surgery. Methods MEDLINE (PubMed®) was searched for literature published between 2010 and 2016. A total of 53 relevant full-text articles were eventually evaluated. Results Ultrasound (US) is an established part of algorithms used to diagnose acute appendicitis and has already replaced CT as the imaging technique of choice. The differences between SPUS and RPUS with regard to sensitivity, specificity, and positive and negative predictive values (PPV, NPV) are not statistically significant. The benefit of SPUS over RPUS is the simultaneous clinical assessment of the patient by the surgeon while the sonogram is performed (sonopalpation), which can increase diagnostic accuracy even further. Radiation exposure as a result of CT could be avoided or significantly reduced through the routine use of US, which is increasingly being used and is widely available. Conclusions SPUS should be the first imaging technique used to diagnose patients with suspected appendicitis. Additional diagnosis using CT or MRI is only recommended if sonographic imaging of the appendix is impossible in combination with specific clinical and laboratory criteria. A structured diagnostic approach with obligatory use of (SP)US, as described in the diagnostic algorithm, should be used for detection of acute appendicitis.

EMMPRIN expression is involved in the development of interstitial fibrosis and tubular atrophy in human kidney allografts.

BACKGROUND: Matrix metalloproteinases (MMP) are involved in the development of interstitial fibrosis and tubular atrophy (IF/TA) in renal disease. The synthesis of MMP is activated by the extracellular matrix metalloproteinases inducer protein (EMMPRIN). To analyze the role of EMMPRIN in IF/TA, we retrospectively detected EMMPRIN expression in specimens of human renal allografts with various levels of IF/TA. METHODS: Immunohistochemistry was performed to detect EMMPRIN expression. In a retrospective analysis, a total cohort of 50 specimens were divided according to BANFF-classification into four subgroups (0-3): no, mild (≤ 25%), moderate (26-50%), or severe (>50%) IF/TA. Among other parameters, renal function was analyzed and compared to EMMPRIN expression. RESULTS: In 24 of 38 biopsies, we detected positive EMMPRIN staining. All nephrectomy (n = 12) samples were negative for EMMPRIN. Positive staining in the biopsy samples was detectable on the basolateral side of tubular epithelial cells. EMMPRIN staining was negatively correlated with IF/TA (p < 0.001). We found significant differences between the mean EMMPRIN expression in IF/TA groups 0 and 3 (p = 0.021) and groups 1 and 3 (p = 0.004). Furthermore, we found significant correlations between EMMPRIN staining and renal function. CONCLUSION: Our data suggest that EMMPRIN is involved in the pathophysiology of IF/TA.

Factors Governing the Precision of Subvisible Particle Measurement Methods - A Case Study with a Low-Concentration Therapeutic Protein Product in a Prefilled Syringe.

PURPOSE: The current study was performed to assess the precision of the principal subvisible particle measurement methods available today. Special attention was given to identifying the sources of error and the factors governing analytical performance. METHODS: The performance of individual techniques was evaluated using a commercial biologic drug product in a prefilled syringe container. In control experiments, latex spheres were used as standards and instrument calibration suspensions. RESULTS: The results reported in this manuscript clearly demonstrated that the particle measurement techniques operating in the submicrometer range have much lower precision than the micrometer size-range methods. It was established that the main factor governing the relatively poor precision of submicrometer methods in general and inherently, is their low sampling volume and the corresponding large extrapolation factors for calculating final results. CONCLUSIONS: The variety of new methods for submicrometer particle analysis may in the future support product characterization; however, the performance of the existing methods does not yet allow for their use in routine practice and quality control.

Measuring Subvisible Particles in Protein Formulations Using a Modified Light Obscuration Sensor with Improved Detection Capabilities.

Although light obscuration is the "gold standard" for subvisible particle measurements in biopharmaceutical products, the current technology has limitations with respect to the detection of translucent proteinaceous particles and particles of sizes smaller and around 2 µm. Here, we describe the evaluation of a modified light obscuration sensor utilizing a novel measuring mode. Whereas standard light obscuration methodology monitors the height (amplitude) of the signal, the new approach monitors its length (width). Experimental evaluation demonstrated that this new detection mode leads to improved detection of subvisible particles of sizes smaller than 2 µm, reduction of artifacts during measurements especially of low concentrations of translucent protein particles, and higher counting accuracy as compared to flow imaging microscopy and standard light obscuration measurements.

Comparative Evaluation of Two Methods for Preparative Fractionation of Proteinaceous Subvisible Particles--Differential Centrifugation and FACS.

PURPOSE: The goal of this study was to compare and evaluate two preparative techniques for fractionation of proteinaceous subvisible particles. This work enables future studies to address the potential biological consequences of proteinaceous subvisible particles in protein therapeutic products. METHODS: Particles were generated by heat stress and separated by size using differential centrifugation and FACS (Fluorescence-activated cell sorter). Resulting fractions were characterized by size-exclusion chromatography, light obscuration, flow imaging microscopy and resonant mass measurement. RESULTS: Here we report the optimization and comprehensive evaluation of two methods for preparative fractionation of subvisible proteinaceous particles into distinct size fractions in the range between 0.25 and 100 µm: differential centrifugation and FACS. Using these methods, well-defined size fractions were prepared and characterized in detail. Critical assessment and comparison of the two techniques demonstrated their complementarity and for the first time-their relative advantages and drawbacks. CONCLUSIONS: FACS and differential centrifugation are valuable tools to prepare well-defined size-fractions of subvisible proteinaceous particles. Both techniques possess unique and advantageous attributes and will likely find complementary application in future research on the biological consequences of proteinaceous subvisible particles.

Subvisible (2-100 µm) particle analysis during biotherapeutic drug product development: Part 2, experience with the application of subvisible particle analysis.

Measurement and characterization of subvisible particles (including proteinaceous and non-proteinaceous particulate matter) is an important aspect of the pharmaceutical development process for biotherapeutics. Health authorities have increased expectations for subvisible particle data beyond criteria specified in the pharmacopeia and covering a wider size range. In addition, subvisible particle data is being requested for samples exposed to various stress conditions and to support process/product changes. Consequently, subvisible particle analysis has expanded beyond routine testing of finished dosage forms using traditional compendial methods. Over the past decade, advances have been made in the detection and understanding of subvisible particle formation. This article presents industry case studies to illustrate the implementation of strategies for subvisible particle analysis as a characterization tool to assess the nature of the particulate matter and applications in drug product development, stability studies and post-marketing changes.

Comparative prognostic value of C-reactive protein & fibrinogen in patients with coronary artery disease.

BACKGROUND & OBJECTIVES: The comparative prognostic value of C-reactive protein (CRP) and fibrinogen for cardiovascular events has been inconclusively investigated. t0 his study was carried out to compare the prognostic value of CRP versus fibrinogen in patients with coronary artery disease (CAD). METHODS: The study included 13,100 patients with coronary angiography-confirmed CAD. Plasma CRP and fibrinogen levels were measured before angiography in all patients. The levels of CRP>3 mg/l and fibrinogen>350 mg/dl were considered as elevated. The primary outcome was 1-year all-cause mortality. RESULTS: Patients were divided into four groups: patients with CRP≤3 mg/l and fibrinogen ≤350 mg/dl (n=4206); patients with CRP≤3 mg/l and fibrinogen >350 mg/dl (n=3132); patients with CRP>3 mg/l and fibrinogen ≤ 350 mg/dl (n=1273) and CRP >3 mg/l and patients with fibrinogen >350 mg/dl (n=4489). There were 634 deaths: 75 deaths in patients with CRP ≤3 mg/l and fibrinogen ≤350 mg/dl, 91 deaths in patients with CRP ≤3 mg/l and fibrinogen >350 mg/dl, 87 deaths in patients with CRP >3 mg/l and fibrinogen ≤350 mg/dl and 381 deaths in patients with CRP >3 mg/l and fibrinogen >350 mg/dl (Kaplan-Meier estimates of all-cause mortality, 1.8, 3.0, 7.0 and 8.7 %, log-rank test P<0.001). The multivariate analysis showed that CRP [adjusted hazard ratio (HR)=1.31, 95% confidence interval (CI) 1.18-1.45, for each standard deviation increase in the logarithmic scale] but not fibrinogen [adjusted HR=0.99 (0.90-1.09), for each standard deviation increase in the logarithmic scale] was an independent correlate of mortality. INTERPRETATION & CONCLUSIONS: The findings indicated that in patients with CAD, CRP was a better predictor of mortality than fibrinogen and offered prognostic information beyond that provided by the conventional cardiovascular risk factors.

Comparison of vascular closure devices vs manual compression after femoral artery puncture: the ISAR-CLOSURE randomized clinical trial.

IMPORTANCE: The role of vascular closure devices (VCD) for the achievement of hemostasis in patients undergoing transfemoral coronary angiography remains controversial. OBJECTIVE: To compare outcomes with the use of 2 hemostasis strategies after diagnostic coronary angiography performed via transfemoral access-a VCD-based strategy with 2 types of devices, an intravascular device and an extravascular device, vs standard manual compression. The primary hypothesis to be tested was that femoral hemostasis achieved through VCD is noninferior to manual compression in terms of vascular access-site complications. A secondary objective was the comparison of the 2 types of VCD. DESIGN, SETTING, AND PARTICIPANTS: Randomized, large-scale, multicenter, open-label clinical trial. We enrolled 4524 patients undergoing coronary angiography with a 6 French sheath via the common femoral artery from April 2011 through May 2014 in 4 centers in Germany. Last 30-day follow-up was performed in July 2014. INTERVENTIONS: After angiography of the access site, patients were randomized to hemostasis with an intravascular VCD, extravascular VCD, or manual compression in a 1:1:1 ratio. MAIN OUTCOMES AND MEASURES: Primary end point: the composite of access site-related vascular complications at 30 days after randomization with a 2% noninferiority margin. Secondary end points: time to hemostasis, repeat manual compression, and VCD failure. An α-level of .025 was chosen for primary and secondary comparisons. RESULTS: Of the 4524 enrolled patients, 3015 were randomly assigned to a VCD group (1509 received intravascular VCD and 1506 received extravascular VCD) and 1509 patients were randomly assigned to the manual compression group. Before hospital discharge, duplex sonography of the access site was performed in 4231 (94%) patients. The primary end point was observed in 208 patients (6.9%) assigned to receive a VCD and 119 patients (7.9%) assigned to manual compression (difference, -1.0% [1-sided 97.5% CI, 0.7%]; P for noninferiority<.001). Time to hemostasis was significantly shorter in patients with VCD (1 minute [interquartile range {IQR}, 0.5-2.0]), vs manual compression (10 minutes [IQR, 10-15]; P < .001). Time to hemostasis was significantly shorter among patients with intravascular VCD (0.5 minute [IQR, 0.2-1.0]), vs extravascular VCD (2.0 minutes [IQR, 1.0-2.0]; P <.001) and closure device failure was also significantly lower among those with intravascular vs extravascular VCD (80 patients [5.3%], vs 184 patients [12.2%]; P < .001). CONCLUSIONS AND RELEVANCE: In patients undergoing transfemoral coronary angiography, VCDs were noninferior to manual compression in terms of vascular access-site complications and reduced time to hemostasis. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01389375.

[Current mortality from war injuries-A narrative review]. / Aktuelle Mortalität von Kriegsverletzungen ­ eine narrative Übersichtsarbeit.

BACKGROUND: The war in Ukraine has led to a strategic reorientation of the German Armed Forces towards national and alliance defense. This has also raised the need for medical and surgical adaptation to scenarios of conventional warfare. In order to develop appropriate and effective concepts it is necessary to identify those war injuries that are associated with a relevant primary and secondary mortality and that can be influenced by medical measures (potentially survivable injuries). OBJECTIVE: The aim of this selective literature review was to identify war injuries with high primary and secondary mortality. METHODS: A selective literature review was performed in the PubMed® database with the search terms war OR combat AND injury AND mortality from 2001 to 2023. Studies including data of war injuries and associated mortality were included. RESULTS: A total of 33 studies were included in the analysis. Severe traumatic brain injury and thoracoabdominal hemorrhage were the main contributors to primary mortality. Injuries to the trunk, neck, traumatic brain injury, and burns were associated with relevant secondary mortality. Among potentially survivable injuries, thoracoabdominal hemorrhage accounted for the largest proportion. Prehospital blood transfusions and short transport times significantly reduced war-associated mortality. CONCLUSION: Control of thoracoabdominal hemorrhage has the highest potential to reduce mortality in modern warfare. Besides that, treatment of traumatic brain injury, burns and neck injuries has a high relevance in reducing mortality. Hospitals of the German Armed Forces need to focus on these requirements.

Outcome of Laparoscopic Versus Open Appendectomy for Acute Appendicitis-Results of a Propensity Score Matching Analysis of 542 Patients and Consequences for the Military Surgeon.

INTRODUCTION: Appendectomy is the treatment of choice for acute appendicitis. The procedure can be performed open or laparoscopically. However, laparoscopy is not available on military missions abroad. It is unclear whether treatment outcomes differ between the two surgical approaches. MATERIALS AND METHODS: Treatment data of all patients undergoing open and laparoscopic appendectomies in the German Armed Forces Hospital of Ulm from 2013 to 2017 were collected retrospectively. A propensity score matching analysis was performed to minimize the influence of potential confounders and to assess the influence of surgical approach on outcome (reoperations, superficial and deep surgical infections, length of postoperative ileus, need for intravenous analgesics, and operative time). RESULTS: A total of 542 patients with complete datasets were included in the propensity score matching analysis, among these 64 with open and 478 laparoscopic procedures. There were no statistically significant differences between open and laparoscopic surgeries with respect to all outcome variables, with the exception of a 25-minute prolonged operative time for the open approach. CONCLUSIONS: Open appendectomy can be considered equivalent in outcome to the laparoscopic procedure and thus can be performed on military missions abroad without compromising outcome. Military surgeons must continue to be trained and confident in open appendectomy.

Diagnostic options for blunt abdominal trauma.

PURPOSE: Physical examination, laboratory tests, ultrasound, conventional radiography, multislice computed tomography (MSCT), and diagnostic laparoscopy are used for diagnosing blunt abdominal trauma. In this article, we investigate and evaluate the usefulness and limitations of various diagnostic modalities on the basis of a comprehensive review of the literature. METHODS: We searched commonly used databases in order to obtain information about the aforementioned diagnostic modalities. Relevant articles were included in the literature review. On the basis of the results of our comprehensive analysis of the literature and a current case, we offer a diagnostic algorithm. RESULTS: A total of 86 studies were included in the review. Ecchymosis of the abdominal wall (seat belt sign) is a clinical sign that has a high predictive value. Laboratory values such as those for haematocrit, haemoglobin, base excess or deficit, and international normalised ratio (INR) are prognostic parameters that are useful in guiding therapy. Extended focused assessment with sonography for trauma (eFAST) has become a well established component of the trauma room algorithm but is of limited usefulness in the diagnosis of blunt abdominal trauma. Compared with all other diagnostic modalities, MSCT has the highest sensitivity and specificity. Diagnostic laparoscopy is an invasive technique that may also serve as a therapeutic tool and is particularly suited for haemodynamically stable patients with suspected hollow viscus injuries. CONCLUSIONS: MSCT is the gold standard diagnostic modality for blunt abdominal trauma because of its high sensitivity and specificity in detecting relevant intra-abdominal injuries. In many cases, however, clinical, laboratory and imaging findings must be interpreted jointly for an adequate evaluation of a patient's injuries and for treatment planning since these data supplement and complement one another. Patients with blunt abdominal trauma should be admitted for clinical observation over a minimum period of 24 h since there is no investigation that can reliably rule out intra-abdominal injuries.

A renewed rise in global HCFC-141b emissions between 2017-2021.

Global emissions of the ozone-depleting gas HCFC-141b (1,1-dichloro-1-fluoroethane, CH3CCl2F) derived from measurements of atmospheric mole fractions increased between 2017 and 2021 despite a fall in reported production and consumption of HCFC-141b for dispersive uses. HCFC-141b is a controlled substance under the Montreal Protocol, and its phase-out is currently underway, after a peak in reported consumption and production in developing (Article 5) countries in 2013. If reported production and consumption are correct, our study suggests that the 2017-2021 rise is due to an increase in emissions from the bank when appliances containing HCFC-141b reach the end of their life, or from production of HCFC-141b not reported for dispersive uses. Regional emissions have been estimated between 2017-2020 for all regions where measurements have sufficient sensitivity to emissions. This includes the regions of northwestern Europe, east Asia, the United States and Australia, where emissions decreased by a total of 2.3 ± 4.6 Ggyr-1, compared to a mean global increase of 3.0 ± 1.2 Ggyr-1 over the same period. Collectively these regions only account for around 30% of global emissions in 2020. We are not able to pinpoint the source regions or specific activities responsible for the recent global emission rise.

Preparation of metallochelating microbubbles and study on their site-specific interaction with rGFP-HisTag as a model protein.

The histidine-metallochelating lipid complex is one of the smallest high affinity binding units used as tools for rapid noncovalent binding of histidine tagged molecules, especially recombinant proteins. The advantage of metallochelating complex over protein-ligand complexes (e.g., streptavidine-biotin, glutathiontransferase-glutathion) consists in its very low immunogenicity, if any. This concept for the construction of surface-modified metallochelating microbubbles was proved with recombinant green fluorescent protein (rGFP) containing 6His-tag. This protein is easy to be detected by various fluorescence techniques as flow cytometry and confocal microscopy. Microbubbles (MB) composed of DPPC with various contents of metallochelating lipid DOGS-NTA-Ni were prepared by intensive shaking of the liposome suspension under the atmosphere of sulfur hexafluoride. For this purpose, the instrument 3M ESPE CapMix was used. Various techniques (static light scattering, flow cytometry, and optical microscopy) were compared and used for the measurements of the size distribution of MB. All three methods demonstrated that the prepared MB were homogeneous in their size, and the mean diameter of the MB in various batches was within the range of 2.1-2.8 µm (the size range of 1-10 µm). The presence of large MB (8-10 µm) was marginal. Counting of MB revealed that the average amount of MB prepared of 10 mg of phospholipid equaled approximately 10(9) MB/mL. Lyophilized MB were prepared with saccharose as a cryoprotectant. These MB were shown to be stable both in vitro (the estimated half-live of the MB in bovine serum at 37 °C was 3-7 min) and in vivo (mouse). The stability of the MB was affected by molar content of DOGS-NTA-Ni. DPPC-based metallochelating MB provided a clear and very contrast image of the ventricular cavity soon after the injection. Site selective and stable binding of rGFP-HisTag (as a model of His-tagged protein) onto the surface of metallochelating MB was demonstrated by confocal microscopy.

Time course of acetylcholine-induced activation of sympathetic efferents matches axon reflex sweating in humans.

Action potentials from postganglionic C-fibres were recorded in healthy volunteers by microneurography in the peroneal nerve. Their responsiveness to mechanical or heat stimuli or to sympathetic reflex provocation tests was determined by transient slowing of conduction velocity following activation. Twenty units were classified as sympathetic efferent units. Acetylcholine (ACh) iontophoresis (10%, 1 mA, 1 min) inside their innervation territory activated 8 of 20 sympathetic fibres with a mean delay of 61 ± 12 s, peak response at 175 ± 38 s, and a duration of 240 ± 42 s, whereas iontophoresis of saline did not activate any of them. The time course of neuronal activation correlated with the axon reflex sweating measured by an evaporimeter in a separate session (delay 76 ± 9 s, peak at 195 ± 12 s, decline to 50% of peak 312 ± 25 s). No ACh-induced vasoconstriction was observed by laser Doppler scanning (n = 11) even after depletion of neuropeptides by chronic topical capsaicin treatment (n = 8). We conclude that ACh iontophoresis activates about half of the sympathetic fibres in human skin and provokes a corresponding axon reflex sweating. The absence of ACh-induced vasoconstriction even after the depletion of neuropeptides by capsaicin suggests that only sudomotor fibres, but not sympathetic vasoconstrictor fibres are activated by this stimulus.