Experience with the use of siltuximab in patients with SARS-CoV-2 infection.

OBJECTIVE: The study aims to describe characteristics and clinical outcome of patients with SARS-CoV-2 infection that received siltuximab according to a protocol that aimed to early block the activity of IL-6 to avoid the progression of the inflammatory flare. METHODS: Retrospective review of the first 31 patients with SARS-CoV-2 treated with siltuximab, in Hospital Clinic of Barcelona or Hospital Universitario Salamanca, from March to April 2020 with positive polymerase-chain reaction (PCR) from a nasopharyngeal swab. RESULTS: The cohort included 31 cases that received siltuximab with a median (IQR) age of 62 (56-71) and 71% were males. The most frequent comorbidity was hypertension (48%). The median dose of siltuximab was 800 mg ranging between 785 and 900 mg. 7 patients received siltuximab as a salvage therapy after one dose of tocilizumab. At the end of the study, a total of 26 (83.9) patients had been discharged alive and the mortality rate was 16.1% but only 1 out of 24 that received siltuximab as a first line option (4%). CONCLUSIONS: Siltuximab is a well-tolerated alternative to tocilizumab when administered as a first line option in patients with COVID-19 pneumonia within the first 10 days from symptoms onset and high C-reactive protein.

Tocilizumab reduces the risk of ICU admission and mortality in patients with SARS-CoV-2 infection.

OBJECTIVE: In some patients the immune response triggered by SARS-CoV-2 is unbalanced, presenting an acute respiratory distress syndrome which in many cases requires intensive care unit (ICU) admission. The limitation of ICU beds has been one of the major burdens in the management around the world; therefore, clinical strategies to avoid ICU admission are needed. We aimed to describe the influence of tocilizumab on the need of transfer to ICU or death in non-critically ill patients. METHODS: A retrospective study of 171 patients with SARS-CoV-2 infection that did not qualify as requiring transfer to ICU during the first 24h after admission to a conventional ward, were included. The criteria to receive tocilizumab was radiological impairment, oxygen demand or an increasing of inflammatory parameters, however, the ultimate decision was left to the attending physician judgement. The primary outcome was the need of ICU admission or death whichever came first. RESULTS: A total of 77 patients received tocilizumab and 94 did not. The tocilizumab group had less ICU admissions (10.3% vs. 27.6%, P=0.005) and need of invasive ventilation (0 vs 13.8%, P=0.001). In the multivariable analysis, tocilizumab remained as a protective variable (OR: 0.03, CI 95%: 0.007-0.1, P=0.0001) of ICU admission or death. CONCLUSIONS: Tocilizumab in early stages of the inflammatory flare could reduce an important number of ICU admissions and mechanical ventilation. The mortality rate of 10.3% among patients receiving tocilizumab appears to be lower than other reports. This is a non-randomized study and the results should be interpreted with caution.

The effect of oblique-angle sputtering on large area deposition: a unidirectional ultrathin Au plasmonic film growth design.

Growing ultrathin nanogranular (NG) metallic films with continuously varying thickness is of great interest for studying regions of criticality and scaling behaviors in the vicinity of quantum phase transitions. In the present work, an ultrathin gold plasmonic NG film was grown on a sapphire substrate by RF magnetron sputtering with an intentional deposition gradient to create a linearly variable thickness ranging from 5 to 13 nm. The aim is to accurately study the electronic phase transition from the quantum tunneling regime to the metallic conduction one. The film structural characterization was performed by means of high-resolution transmission electron microscopy, atomic force microscopy, as well as x-ray diffraction and reflectivity techniques, which indicate the Volmer-Weber film growth mode. The optical and electrical measurements show a transition from dielectric-isolated gold NPs towards a continuous metallic network when t becomes larger than a critical value of tM = 7.8 nm. Our results show that the onset of the percolation region occurs when a localized surface plasma resonance transforms to display a Drude component, indicative of free charge carriers. We demonstrate that, by using a continuously varying thickness, criteria for metallicity can be unambiguously identified. The onset of metallicity is clearly distinguished by the Drude damping factor and by discontinuities in the plasma frequencies as functions of thickness.

An optical-based biosensor of the epithelial sodium channel as a tool for diagnosing hypertension.

Arterial hypertension (HTN) is a world health concern presenting difficulties for its early detection. It leads to cardiovascular and kidney complications that increase morbidity in adults. Overexpression in the epithelial sodium channel (ENaC) in membrane platelets can be related with the presence of HTN and thus can be used as a biomarker to detect this medical condition. Here, we propose a method for HTN diagnosis based on gold nanoparticles (GNPs) conjugated to an antibody against the ENaC present on platelets. These functionalized GNPs were analyzed by Zeta potential, dynamic light scattering, electron microscopy, and other spectroscopic techniques. To verify that the GNPs and α-ENaC antibodies formed conjugates (GNPs-antiENaC) that maintained their specificity to the target, we carried out an indirect immunofluorescence detection assay of GNPs-antiENaC bound to a secondary antibody labeled with a fluorophore. Our results show that the presence of GNPs increase the fluorescence intensity in platelets treated with GNPs-antiENaC conjugates. It is also observed a clear tendency of the fluorescence signal in platelets treated with the conjugates that could be used for discrimination between normotensive and hypertensive samples. The proposed assay can be implemented as a very sensitive routine test to diagnose HTN.

Alterations to the contents of plasma membrane structural lipids are associated with structural changes and compartmentalization in platelets in hypertension.

Arterial hypertension (HTN) can lead to serious organ damage. Several mechanisms have been implicated in the pathogenesis of HTN including constitutive activation of platelets, which increases the risk of aggregation and clot formation. We recently demonstrated the plasma membranes of platelets from patients with HTN exhibit modified structural and physicochemical properties; Raman and Fourier transform infrared by attenuated total reflectance (FTIR-ATR) spectroscopy also indicated lipid content and protein structure alterations. This study aimed to precisely quantify the constituents of the main structural phospholipids and cholesterol in the plasma membranes of platelets from patients with HTN and normotensive individuals. We also assessed the consequence of these alterations on platelet structure and function. Liquid chromatography coupled to triple quadrupole mass spectrometry revealed the plasma membranes of HTN platelets contained less cholesterol and phosphatidylcholine, more phosphatidylserine and phosphatidylethanolamine and had similar sphingosine contents. Atomic force microscopy revealed HTN platelets exhibited increased surface roughness and more pleats. Transmission electron microscopy revealed diminution of the internal membranous structures in HTN platelets. Our findings strongly suggest plasma membrane lipid content alterations-including cholesterol depletion-occur in HTN, and these alterations may induce morphological and physiological abnormalities that participate in the functional changes associated with hypertension.

The bifoil photodyne: a photonic crystal oscillator.

Optical tweezers is an example how to use light to generate a physical force. They have been used to levitate viruses, bacteria, cells, and sub cellular organisms. Nonetheless it would be beneficial to use such force to develop a new kind of applications. However the radiation pressure usually is small to think in moving larger objects. Currently, there is some research investigating novel photonic working principles to generate a higher force. Here, we studied theoretically and experimentally the induction of electromagnetic forces in one-dimensional photonic crystals when light impinges on the off-axis direction. The photonic structure consists of a micro-cavity like structure formed of two one-dimensional photonic crystals made of free-standing porous silicon, separated by a variable air gap and the working wavelength is 633 nm. We show experimental evidence of this force when the photonic structure is capable of making auto-oscillations and forced-oscillations. We measured peak displacements and velocities ranging from 2 up to 35 microns and 0.4 up to 2.1 mm/s with a power of 13 mW. Recent evidence showed that giant resonant light forces could induce average velocity values of 0.45 mm/s in microspheres embedded in water with 43 mW light power.

Aqueous root extracts from Mimosa albida Humb. & Bonpl. ex Willd display antinociceptive activity in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: In this work, we study whether aqueous extracts from the roots of Mimosa albida Humb. & Bonpl. ex Willd, a plant known in the Highlands of Chiapas, Mexico as "Lotóm chíx" are endowed with both antinociceptive and anxiolytic effects. MATERIALS AND METHODS: ICR mice were systemically treated with aqueous extracts from Mimosa albida and the reference compounds (diazepam, dipyrone and/or fentanyl) and their behavior was evaluated in several behavioral tests. RESULTS: Administration of aqueous extracts from the roots of Mimosa albida resulted in a reduction of the nociception elicited in mice by both the hot plate (12.5, 25 and 50 mg/kg; i.p.) and the acetic acid-induced writhing (25 and 50 mg/kg; i.p.) tests. No effects were however observed both in the elevated plus-maze and hole board test (3.2, 12.5 and 25 mg/kg; i.p.). In contrast, both locomotion (open field test) and motor coordination (rotarod test) were affected at doses (50, 100 y 200 mg/kg; i.p.) higher than those having antinociceptive effects. CONCLUSION: These data suggest that in mice the systemic administration of low doses of aqueous extracts from the roots of Mimosa albida results in antinociceptive effects in several models of pain through mechanisms that do not involve the opioid system pathway. These results support the ethnopharmacological use of Mimosa albida in popular medicine.

Intercalated and paracapsular cell islands of the adult rat amygdala: a combined rapid-Golgi, ultrastructural, and immunohistochemical account.

The anterior and rostral paracapsular intercalated islands (AIC and PIC, respectively) were studied in the context of the amygdaloid modulation of fear/anxiety using horizontal sections. The structural analysis carried out using silver-impregnated specimens revealed that the AIC is composed of tightly packed, medium-sized spiny neurons with distinct dendritic and axonal patterns that send projecting axons to the central nucleus of the amygdala. The AIC occupies a strategic position between the basolateral amygdaloid complex and the caudal limb of the anterior commissure from which it receives fibers en passage and axon terminals. Electron microscopic observation of terminal (i.e., synaptic) degeneration 72 h after the surgical interruption of the anterior commissure, confirms the synaptic interaction between the latter and the AIC neurons. These observations suggest that these islands may gate the activity of neurons from the contralateral basal forebrain and synchronize the anxiogenic output of both amygdalae. Immunohistochemical analysis indicated that, within the AIC and rostral PIC, the distance between tyrosine hydroxylase-immunoreactive terminals and the punctate dopamine D(1) receptor immunoreactivity, was in the micrometer range. These results indicate a short distance and a rapid extrasynaptic form of dopamine volume transmission mediated via D(1) receptors in the AIC and PIC which may enhance fear and anxiety by suppressing feed-forward inhibition in the basolateral and central amygdaloid nuclei. The strong suggestion for a commissural axon projection to the AIC documented here, coupled with the previous evidences indicting an isocortical and amygdalar contributions to the anterior commissure, opens the possibility that the AIC may be involved in decoding nerve impulses arising from both the ipsi- and contra-lateral forebrain to, in turn, modulate the homolateral amygdala.

Distribution of dopamine D(2)-like receptors in the rat amygdala and their role in the modulation of unconditioned fear and anxiety.

Amygdaloid dopamine D(2) receptors play an important role in the modulation of fear/anxiety. Their topographical distribution within the amygdala is however unclear, and their role in unconditioned fear/anxiety remains largely unknown. The aim of this paper was to study the intra-amygdaloid distribution of D(2) receptors and to ascertain their role in unconditioned anxiety. Chemical anatomical studies in the rat, using D(2) and D(3)in situ hybridization, quantitative receptor autoradiography with either [(3)H]raclopride or [(125)I]sulpiride, and D(2)-like immunocytochemistry showed that the highest density of dopamine D(2) receptors is present in the central amygdaloid nucleus, particularly within its latero-capsular division, in which a D(2) but not a D(3) mRNA signal was observed. However, although at considerably reduced densities dopamine D(2) receptors were also found in other locations within the amygdala, including the basolateral nucleus. Behaviorally, the infusion of raclopride (0.75-4 µg/side) in the area of the central amygdaloid nucleus resulted at low doses in the appearance of anxiogenic-like effects in the Shock-Probe Burying test, whereas no effects of raclopride treatment were found at any dose in the Elevated Plus-Maze and the Open-Field test. Our results indicate that amygdaloid dopamine D(2)-like receptors have a topographically differentiated distribution within the rat amygdala, the major location being in the central amygdaloid nucleus. D(2)-like receptors play a role in the modulation of anxiety responses involving a potential differential function of D(2)-like receptors in the central amygdaloid nucleus versus the basolateral amygdaloid nucleus.

Anomalous patterned scattering spectra of one-dimensional porous silicon photonic crystals.

Far-field secondary emission spectra of one-dimensional periodic photonic structures based on porous silicon show characteristic co-focal rings centered close to the structure plane normal. The rings appear when the frequency of picosecond excitation laser pulses is tuned to the edges of the fourth photonic band gap. They can be clearly distinguished from the typical reflected and transmitted light in the oblique incidence geometry. The rings number is dependent on the excitation frequency and the incidence angle. We explain these anomalous spectral features of porous silicon structures by the spectral filtering of light elastically scattered inside the photonic structure by the narrow photonic bands. The elastic scattering of light due to the photonic disorder in the structure causes the appearance of secondary waves propagating in any direction. But only those waves which fall into the allowed photonic bands penetrate through the whole structure and move through its front or back surfaces. The observed patterned secondary emission is an example of efficient photonic engineering by simple means of multilayer porous silicon structures.

Molecular and catalytic properties of the aldehyde dehydrogenase of Gluconacetobacter diazotrophicus, a quinoheme protein containing pyrroloquinoline quinone, cytochrome b, and cytochrome c.

Several aldehyde dehydrogenase (ALDH) complexes have been purified from the membranes of acetic acid bacteria. The enzyme structures and the chemical nature of the prosthetic groups associated with these enzymes remain a matter of debate. We report here on the molecular and catalytic properties of the membrane-bound ALDH complex of the diazotrophic bacterium Gluconacetobacter diazotrophicus. The purified ALDH complex is a heterodimer comprising two subunits of 79.7 and 50 kDa, respectively. Reversed-phase high-pressure liquid chromatography (HPLC) and electron paramagnetic resonance spectroscopy led us to demonstrate, for the first time, the unequivocal presence of a pyrroloquinoline quinone prosthetic group associated with an ALDH complex from acetic acid bacteria. In addition, heme b was detected by UV-visible light (UV-Vis) spectroscopy and confirmed by reversed-phase HPLC. The smaller subunit bears three cytochromes c. Aliphatic aldehydes, but not formaldehyde, were suitable substrates. Using ferricyanide as an electron acceptor, the enzyme showed an optimum pH of 3.5 that shifted to pH 7.0 when phenazine methosulfate plus 2,6-dichlorophenolindophenol were the electron acceptors. Acetaldehyde did not reduce measurable levels of the cytochrome b and c centers; however, the dithionite-reduced hemes were conveniently oxidized by ubiquinone-1; this finding suggests that cytochrome b and the cytochromes c constitute an intramolecular redox sequence that delivers electrons to the membrane ubiquinone.

Role of cholecystokinin-A and cholecystokinin-B receptors in anxiety.

Evidence from several laboratories indicates that the anxiogenic effects of cholecystokinin (CCK) are mediated by CCKB receptors. However, it has been reported that CCKA receptors have been found in brain and CCKA antagonists have anxiolytic properties. The aim of this work was to study whether CCKA receptors are also involved in the modulation of anxiety. Anxiogenic effects were observed in the elevated plus maze in rats when pure CCKB receptor agonists (CCK-4 and CCK-8 non-sulfated) or CCK-8S, a CCKB/CCKA agonist, were injected into the lateral ventricle. In contrast, CCK-33, a CCKA agonist or CCK-(1-21) and CCK-(26-29) were ineffective. Furthermore, the anxiogenic effects of CCK-8S were prevented by blocking CCKB but not CCKA receptors. Finally, CCK-33 injected into the postero-medial nucleus accumbens failed to affect the anxiety level of the rats. These results indicate that CCKA receptors are not involved in anxiety, as measured by the paradigms used in this work.

Lack of coupling between GABA release and GABA synthesis in the rat brain via GABA(B) autoreceptors.

GABA is synthesized within GABA terminals through a highly compartmentalized process in which glial-derived glutamine is a major precursor and its release is modulated by GABA(B) autoreceptors. The aim of this work was to ascertain whether or not GABA synthesis and release are coupled in the rat brain through a GABA(B) autoreceptor-mediated modulation. It was found that (-)baclofen (30microM) reduces the K+ stimulated release of [3H]GABA in synaptosomes and prisms (10microM) from cerebral cortex, while at the same concentrations (-)baclofen failed to modify the synthesis of [3H]GABA from [3H]glutamine in cortical and hypothalamic slices, prisms and in cortical synaptosomes. In this latter preparation, identical results were observed when (-)baclofen was added to Krebs-Tris media, containing 5 or 15 mM K+ concentration. In agreement with these latter results, glutamic acid decarboxylase (GAD) activity from cortical and hypothalamic prisms was not affected by 1-100 microM (-)baclofen. Similar results on GABA synthesis were also observed when 1-100 microM 3-aminopropil(methyl)phosphinic acid or GABA was used instead of (-)baclofen to stimulate GABA(B) autoreceptors. [3H]GABA release, [3H]GABA synthesis from [3H]glutamine and GAD activity were also insensitive to the action of the GABA(B) antagonist CGP 52432 (10-100microM). Likewise, muscimol (0.3-100microM) did not affect GABA synthesis. Our results indicate that unlike GABA release, GABA synthesis is not modulated by GABA(B) autoreceptors.

The vigilance promoting drug modafinil increases extracellular glutamate levels in the medial preoptic area and the posterior hypothalamus of the conscious rat: prevention by local GABAA receptor blockade.

The effects of modafinil on glutamatergic and GABAergic transmission in the rat medial preoptic area (MPA) and posterior hypothalamus (PH), are analysed. Modafinil (30-300 mg/kg) increased glutamate and decreased GABA levels in the MPA and PH. Local perfusion with the GABAA agonist muscimol (10 microM), reduced, while the GABAA antagonist bicuculline (1 microM and 10 microM) increased glutamate levels. The modafinil (100 mg/kg)-induced increase of glutamate levels was antagonized by local perfusion with bicuculline (1 microM). When glutamate levels were increased by the local perfusion with the glutamate uptake inhibitor L-trans-PDC (0.5 mM), modafinil produced an additional enhancement of glutamate levels. Modafinil (1-33 microM) failed to affect [3H]glutamate uptake in hypothalamic synaptosomes and slices. These findings show that modafinil increases glutamate and decreases GABA levels in MPA and PH. The evidence that bicuculline counteracts the modafinil-induced increase of glutamate levels strengthens the evidence for an inhibitory GABA/glutamate interaction in the above regions controlling the sleep-wakefulness cycle.

Effects of the vigilance promoting drug modafinil on the synthesis of GABA and glutamate in slices of rat hypothalamus.

The effects of the vigilance promoting drug modafil were studied ex vivo (100 mg/kg; i.p.) and in vitro (10-1000 microM modafinil) on the synthesis of [3H]gamma-aminobutyric acid ([3H]GABA) and [3H]glutamate from [3H]glutamine within the rat hypothalamus. No effects of modafinil were observed on the overall synthesis of these neurotransmitters nor, in vitro (1-33 microM modafinil) on other parameters related to the compartmentalization of their synthesis (glutamate decarboxylase and phosphate-activated glutaminase activities, and [3H]glutamine uptake). It is suggested on these grounds, that the modafinil-induced reductions and increases in regional GABA and glutamate extracellular levels respectively using in vivo microdialysis may be a consequence of an indirect effect of modafinil on these neurons.

Biochemical effects induced by REM sleep deprivation in naive and in D-amphetamine treated rats.

The neurochemical dysfunction present in patients showing self-mutilating behavior (SMB) is not well understood. In animal models, rapid eye movement (REM) sleep deprivation enhances the SMB induced by the chronic administration of d-amphetamine. To understand the mechanism underlying these effects the levels of dopamine (DA), noradrenaline (NA) and serotonin (5-HT) were measured in REM sleep deprived only, and in REM sleep deprived and d-amphetamine treated rats. DA levels were elevated (31%) after REM sleep deprivation (48 h) in the neostriatum and the cerebral cortex (33%), while the levels of NA and 5-HT remained constant. A 6-day treatment with d-amphetamine (7.5 mg/kg; i.p.) failed to affect, in REM sleep deprived rats, DA, NA and 5-HT levels. It was also found that REM sleep deprivation had no effects on the d-amphetamine induced [3H]DA release from slices of the same regions. Our results suggests that dopaminergic mechanisms are involved in the effects of REM sleep deprivation on SMB.

GABA-dopamine receptor-receptor interactions in neostriatal membranes of the rat.

Recent evidence has shown in membrane preparations that the binding of one ligand to its receptor is able to modify the binding parameters of a second receptor (receptor-receptor interactions), allowing the modulation of incoming signals onto a neuron. To further understand the gamma-amino-butyric acid (GABA)-dopamine (DA) interactions in the neostriatum we have carried out experiments to explore whether an activation of the GABA(A) receptor could affect the binding characteristics of the D2 DA receptor in membrane preparations of the rat neostriatum. The results show the GABA (30-100 nM) significantly increases the dissociation constant of the high affinity (KH) D2 DA binding site (labelled with the selective D2 DA receptor antagonist [3H]raclopride and that such an effect is fully counteracted by the GABA(A) receptor antagonist bicuculline (1 microM). It is suggested that such putative GABA(A)/D2 receptor-receptor interactions may take place in the somato-dendritic membrane of the striato-pallidal GABA neurons and that it may modulate the inhibitory effects of DA on these neurons, mediated via D2 receptors.

Effects of REM sleep deprivation on the d-amphetamine-induced self-mutilating behavior.

It is well known that self-mutilating behavior (SMB) is developed in rats and humans during the daily treatment with d-amphetamine. Accordingly, in this work it was found that the daily treatment with 7.5 mg/kg d-amphetamine induced in rats a progressive appearance of SMB. Lower doses (5.0 mg/kg) were uneffective and higher doses (10 mg/kg) produced a pattern of SMB in which the mutilation induced at the beginning of the d-amphetamine administration disappears completely as the treatment progresses. Interestingly, it was also found that REM sleep deprivation (48 h) potentiated significantly the SMB induced by the daily administration of 7.5 mg/kg d-amphetamine, and to lesser extent, the SMB induced by the daily treatment with 10 mg/kg d-amphetamine. R(+)-SCH-23390 a D1 dopamine (DA) receptor antagonist blocked completely or abolished the SMB induced by 7.5 mg/kg d-amphetamine in REM sleep deprived rats while (+/-)-sulpiride a D2 DA receptor antagonist had only a partial blocking effect. Haloperidol a D1/D2 DA receptor antagonist behaved as a D1 antagonist. Our results indicate that REM sleep deprivation enhances the SMB induced by the daily administration of d-amphetamine and suggest the involvement of D1 DA receptors in the mechanism underlying the SMB. A role of REM sleep deprivation is also suggested in the appearance of self-mutilating episodes in d-amphetamine addicts.

Modafinil and cortical gamma-aminobutyric acid outflow. Modulation by 5-hydroxytryptamine neurotoxins.

The acute or chronic administration of modafinil, (diphenyl-methyl-sulfinyl-2-acetamide, 30 mg/kg s.c.) decreased gamma-amino-butyric acid (GABA) outflow from the cerebral cortex of freely moving guinea pigs and rats. In 5,7-dihydroxytryptamine intracerebroventricularly pretreated guinea pigs, the effect of modafinil on GABA outflow was reversed and the noradrenaline cortical levels increased. Prazosin (35.8 ng/kg i.p.) blocked the drug-induced increase in GABA efflux. In vitro experiments, performed in rat cortical slices, showed that modafinil failed to affect [3H]GABA release and uptake as well as glutamic acid decarboxylase activity. In conclusion, our results suggest that the balance between central noradrenaline and 5-hydroxytryptamine transmission is important for the regulation by modafinil of the GABAergic release in the cerebral cortex.

Free-GABA levels in the cerebrospinal fluid of patients suffering from several neurological diseases Its potential use for the diagnosis of diseases which course with inflammation and tissular necrosis.

Free GABA levels were measured in the cerebrospinal fluid (CSF) of 74 neurological patients suffering from cerebral cysticercosis (n = 9), Parkinson's disease (n = 5), multiple sclerosis (n = 6), epilepsy (n = 24), meningeal tuberculosis (n = 6), viral encephalitis (n = 3), cerebrovascular disease (n = 8) and several kinds of dystonia (n = 5). A statistical significant four-fold elevation in free GABA levels was found in patients with cerebral cysticercosis. A non statistical significant two-fold increase in free GABA levels was also encountered in the CSF of patients affected by cerebrovascular disease and viral encephalitis. No changes in CSF free GABA levels were found in patients suffering from any of the other disorders. It is suggested that free GABA levels may be elevated in the CSF of patients suffering from neurological diseases which course with inflammation and tissular necrosis such as cerebral cysticercosis. Much work is needed however to establishd whether CSF free GABA levels can be used as a diagnostic tool in at least some type of these patients.