Resprouts from slash-and-burn

Resprouts from slash-and-burn find more events enjoy several advantages when competing against most plants starting from seed (Kammesheidt, 1999). The BN resprouts possess a deep and well-developed root system that favors water and nutrient intake (Kainer et al., 1998). Their above-ground growth in full-light conditions helps them cope with the dense and entangled understory of early forest succession. This ability to resprout renders the tree particularly resilient to SC disturbances. A good

indication of the BN tree’s resprouting capability was the ratio of individuals with resprouted versus uncut stems. This ratio was almost four times higher (3.7:1) in sites that had previously experienced two or more slash-and-burn cycles. Most resprouts exhibited Volasertib nmr multiple stems, and the number

of living shoots increased with the number of times that the resprouts survived the SC events (Fig. 2c). Nevertheless, as observed by Kammesheidt (1998) for many species in fallows exposed to SC, the abundance of stems is later reduced by self-thinning. The importance of resprouting as a demographic process depends on the frequency of severe disturbances, the probability that the species will resprout after them, and the rates of survival, growth and maturity of the resprouts (Paciorek et al., 2000). The only reference that we found regarding the maturity of resprouted BN trees reported anecdotal information from forest dwellers (Baider, 2000), who mentioned that resprouted trees die before they reach reproductive

age. Our findings contradict this opinion because the majority of individuals present in fallows assigned to protection were resprouted trees. Although we did not collect data to address this question, the fact that resprouted multi-stem adults are owned and protected by extractivists is a good indication of their productivity. Adult BN trees have very large crowns. Because many mature trees cannot coexist in the check limited space available, the abundance of seedlings and saplings will ultimately be reduced in number through intraspecific competition. Considerations of this sort allow us to deduce a practical limit for the regeneration density increase and, consequently, a sufficient number of SC cycles after which the BN accumulation becomes redundant. In contrast, another landholder choice having decisive impact is the conversion of crops or fallows into pasture. Once this change has taken place, the development of previously established regeneration is no longer feasible, and that particular site will lose its potential to contribute a high-density BN stand.

, 2007) Cre recombinase

, 2007). Cre recombinase Selleckchem CDK inhibitor is widely used in mouse genetics and has been intensively studied ( Glaser et al., 2005 and Van Duyne, 2001). Particularly in clinical applications, it seems to be advantageous that such recombinases, including Tre, neither produce DSBs nor require additional host factors such as the NHEJ pathway. As a result, the recombination process is very precise and usually error-free ( Glaser et al., 2005 and Van Duyne, 2001). Nevertheless, prior to clinical application various potential

problems connected with the Tre technology have to be resolved. For example, current Tre-recombinase was raised against a primary HIV-1 subtype A isolate (Blackard et al., 1999). It is therefore expected that for broader applications a Tre-recombinase also recognizing a majority of HIV-1 subtypes must be developed. Likewise, Tre treatment may select for outgrowth of resistant viruses resulting from target (loxLTR) site mutation. Both aspects may be addressed by identifying Tre target sequences that are highly conserved in the LTRs of a vast majority of HIV-1 isolates. The recent development of a novel “locus of recombination Selleck GSK2118436 site” search tool and the description of a collection of conserved sequences covering a maximum of HIV-1 variants will

certainly be helpful in achieving this goal (McIntyre et al., 2009 and Surendranath et al., 2010). Even if it turns out that a sterilizing cure cannot be achieved, Tre technology may also be applicable in a functional cure for ex vivo treatment of PBMCs. For this, Tre-recombinase could be expressed as a fusion with a cell-penetrating protein transduction domain (PTD) or membrane translocation motif (TLM) ( Fonseca et al., 2009). As reported recently, directly adding recombinant PTD/TLM-Tre fusion protein to a productively

infected T cell culture resulted in efficient protein translocation and excision of the full-length HIV-1 proviral Niclosamide DNAs from their chromosomal integration sites ( Mariyanna et al., 2012). The growing recognition that a cure for HIV infection is not only needed but also feasible is based on significant advances in basic, translational, and clinical research (Deeks et al., 2012). The remarkable case of the “Berlin patient” particularly revived the idea of gene therapy strategies to eradicate HIV (Kiem et al., 2012 and van Lunzen et al., 2011). Indeed, expression of in vitro engineered enzymes disrupting the CCR5 surface receptor and/or excising the HIV-1 proviral DNA may become critical components of future therapies aiming at virus eradication. It is generally expected that, if achievable at all, no single approach will lead to a sterilizing cure. Rather, a clever combination of drug treatments, therapeutic vaccination strategies, possibly in combination with antiviral gene therapy, likely offers the highest hope for defeating HIV.

The data was sampled at a rate of 1000 Hz The data were analyzed

The data was sampled at a rate of 1000 Hz. The data were analyzed online by the experimenter Selleckchem Atezolizumab and if participants did not keep fixation the trial was discarded and repeated. The results are presented in Fig. 3. All data were tested for normality using the Shapiro–Wilk statistic; the data were normal unless otherwise stated. Inferential statistics used a significance level of p < .05, except when multiple comparisons were performed, where a Bonferonni correction of p < .016 was applied. For both tasks less than 1% of trials were redone because participants failed to keep fixation (CBT: 0.58%; Visual Patterns: 0.56%). Analyses are concerned with the mean span for each condition.

A 2 × 2 × 3 repeated measures ANOVA with the factors Task (Visual, Spatial), Side of Presentation (Temporal, Nasal), and Eye Position (Frontal, Protein Tyrosine Kinase inhibitor Abducted 20, Abducted 40) was performed. A significant

main effect of Task was found, F(1,13) = 235.68; p = .00, with memory span being higher in the visual patterns task (M = 7.38, SE = .26) compared to the Corsi Blocks task (M = 4.72; SE = .22); therefore, the two tasks are analyzed separately. The only statistically significant result was the interaction between Task and Side of Presentation, F(1,13) = 6.27; p = .026. A 2 × 3 repeated measures ANOVA with the factors Side of Presentation (Temporal, Nasal), and Eye Position (Frontal, Abducted 20, Abducted 40) revealed no significant main effects (Side of Presentation: p = .625; Eye Position: p = .280). The interaction was also not statistically significant (p = .682, η2 = 0.2). The same 2 × 3 repeated measures ANOVA was performed for Corsi spans. While the main effect

of Eye Position was not statistically significant (p = .145, η2 = 0.14), the main effect of Side of Presentation was, F(1,13) = 11.56; p = .005, η2 = 0.47 with span being higher in the nasal conditions (M = 4.86, SE = .22) compared to the temporal conditions (M = 4.58, SE = .23). The interaction was not significant (p = .393, η2 = 0.069). Bonferroni-corrected planned comparisons (paired samples t-tests; corrected alpha level p < .016) revealed that Corsi span in the temporal hemifield was significantly impaired compared to span in the nasal hemifield, but only in the Abducted 40 condition t(13) = 2.84; p = .014, d = .78; span reduced Sitaxentan by .42 (SE = .15). There was a trend in the same direction in the Abducted 20 condition that did not approach significance when corrected for multiple comparisons (t(13) = 2.12; p = .053; d = .59). There was no difference in performance in the Frontal condition condition t(13) = .89; p = .39, d = .23). Memory span on the Corsi Blocks task was significantly reduced only when presented locations could not be encoded as the goal of saccadic eye movements; i.e., when memoranda were presented in the temporal hemifield in the 40° eye-abducted condition.

The primary goal of glaucoma treatment is to reduce intraocular p

The primary goal of glaucoma treatment is to reduce intraocular pressure (IOP) using antiglaucoma eye drops, laser treatment, or surgery [2] and [3]. Antiglaucoma eye-drop application is the most common therapy, and can significantly lower IOP and delay glaucoma progression Rapamycin ic50 [4] and [5]. However, patients with glaucoma who use antiglaucoma eye drops have been shown to have a higher prevalence of ocular surface disease than the normal population [6] and [7].

Irritation and conjunctival hyperemia induced by dry eyes are among the main problems when treating patients with glaucoma who require a lifetime management [8], [9] and [10]. Dry-eye therapy has been solely symptomatic, mainly by the application of artificial tears. However, numerous recent studies have demonstrated that inflammation and apoptosis may play key roles in the development

of dry eye syndrome (DES) [11], [12], [13], [14], [15] and [16]. Ginseng (the root of Panax ginseng Meyer) is a valuable folk medicine used in East Asian countries. The two kinds of ginseng, air-dried white ginseng and steamed red ginseng, harbor a variety of active components, including ginsenosides, polysaccharides, peptides, polyacetylenic alcohols, and fatty acids, and its diverse pharmacological effects have been observed in the central nervous system Z-VAD-FMK in vitro and the cardiovascular, endocrine, and immune systems [17], [18], [19], [20], [21], [22], [23], [24] and [25]. Ponatinib solubility dmso Korean Red Ginseng

(KRG) is known to have more pharmacological effects than raw ginseng because of the change of its chemical components (such as Rg3 and Rh2) that are produced in the steaming process [26]. Because of chronic inflammation, conjunctival pathological changes, including squamous metaplasia and goblet cell loss, have been found on cytological analysis of dry eye disease and, thus, anti-inflammatory drugs, such as topical steroid and cyclosporine A, are effective agents for DES [27] and [28]. In an earlier study performed by the authors [29], participants stated that the discomfort caused by antiglaucoma eye drops was relieved by KRG intake. Furthermore, the symptoms and signs of dry eyes were improved in some of these patients. In this randomized, double-blind, placebo-controlled study, we examined the effect of KRG supplementation on DES in patients with glaucoma. This prospective, randomized, double-blind, placebo-controlled, parallel group study was performed at the glaucoma clinic of the Severance Hospital, Seoul, Korea. The study was conducted in accordance with the Declaration of Helsinki, and informed written consent was obtained from each participant. The Institutional Review Board of the Yonsei University Health System approved the study protocol. Participants were enrolled prospectively between July 2013 and December 2013.

They are only likely to be effaced by igneous or high-grade metam

They are only likely to be effaced by igneous or high-grade metamorphic processes, or by erosion once they reach the surface. As with shallow and surface phenomena, anthroturbation fabrics will reach the surface if the crust is eroded following tectonic uplift. Uplift and denudation rates vary considerably, depending on the tectonic setting, but typically do not exceed a couple of millimetres a year (e.g. Abbott et al., 1997 and Schlunegger and Hinderer, 2002); structures a few kilometres

deep will not break the surface for millions to tens of millions of years. Structures on currently stable or descending crust may of course remain preserved below the surface for very much longer, or even permanently. The expression of deep mines and boreholes (particularly once they reach the surface, in

the far geological future) will differ. selleck products Mines – particularly those, such as coalmines that exploit stratabound minerals – will show stratigraphically-related patterns of occurrence. Thus, in each of many coal-fields, that today have substantial outcrops and subcrops in many parts of the world (Fig. 2 for the UK), there can be up to several tens of coal seams exploited to depths that may exceed a kilometre. Each of these seams, over that lateral and vertical extent, will be largely replaced by a horizon marked by little or no remnant coal, but considerable brecciation of adjacent strata (while fossilized examples of, say pit props or mining machinery (or the skeletons of pit ponies or even miners) might occasionally be encountered). In between these intensely worked units there will be thick successions of overlying and underlying strata that are effectively pristine, other than being penetrated in a few places by access shafts and exploration boreholes. Boreholes into present-day oilfields are abundant globally (the total length of oil

boreholes), the great majority drilled since the mid-20th century, has been estimated at 50 million km (J.P.M. Syvitski, personal communication), roughly equivalent to the many length of the present-day global road network or the distance from the Earth to Mars. For each human on Earth today there is thus a length of oil borehole of some seven metres – their share (on average) in the provision of the liquid energy that helps shape their lives. The density of boreholes in oilfields may be seen, for instance, in the map showing the 50,686 wells drilled to date in American waters of the Gulf of Mexico (see Boreholes are structures that in reality penetrate long crustal successions. However, once exhumed in the far future, they may only rarely be encountered in typical rock exposures as lengths of (usually) vertical disruption at decimetre to metre scale in width.

Finally, we discuss the percentage of these 178 subjects that pre

Finally, we discuss the percentage of these 178 subjects that presented with an increase of eosinophils in peripheral blood and BALF. A 70-year-old woman, who has had a diagnosis of sarcoidosis since she was 38 years old, was admitted to our hospital in October 2008.

She had been followed without treatment after the initial diagnosis, but she developed cough and dyspnea in September 2008. She had never smoked Selleck Adriamycin and had no apparent history of exposure to allergens, had not been exposed to environmental changes, and had not used new medicines or dietary supplements for at least 6 months before admission. She had no history of any allergic diseases, such as bronchial asthma, allergic rhinitis, or atopic dermatitis. She lived in an urban area and had never traveled to areas with a high prevalence of parasites. Laboratory data revealed increases in peripheral eosinophil counts (10.0%, 869/μL), total serum IgE levels [509.3 IU/L (normal: 0–295)],

serum angiotensin converting enzyme (ACE) activity [33.5 IU/L (normal: 8.3–21.5)], and soluble interleukin-2 receptor (sIL2R) [3932 U/mL (normal: 0–459)] relative to data from three months before (eosinophil: 5.0%, 330/μL, ACE: 23.2 IU/L). The results of IgE multiple antigen simultaneous tests revealed positive for Dermatophagoides farinae and negative for Aspergillus. She was also negative for antineutrophil cytoplasmic antibodies to both myeloperoxidase and proteinase 3. No evidence of parasites were found in stool samples. High-resolution computed tomography (CT) revealed diffuse ground-glass opacity in the bilateral middle and lower lobe, and enlargement of bilateral JAK inhibitor hilar and mediastinal lymph nodes. Analysis of BALF obtained from the right S4+5 segment

revealed elevated eosinophil (15.2%) and lymphocyte (51.2%) counts and a high CD4/8 ratio (26.2), but no evidence of microbial culture. Lung biopsy revealed infiltration of eosinophils in alveolar walls and alveolar spaces ( Fig 1a) with granulomatous tissue containing multinucleated giant cells ( Fig 1b), but no evidence of angiitis. Transbronchial needle biopsy from the #7 lymph node also revealed non-caseating epithelioid cell granuloma ( Fig. 1c) with no apparent pathogens, such as fungus or mycobacterium. Eosinophilic pneumonia accompanied by sarcoidosis was suspected. Her respiratory symptoms, 4-Aminobutyrate aminotransferase the ground-glass opacities by chest X-ray and peripheral eosinophilia, elevated serum ACE activity and sIL2R remitted spontaneously without treatment within several months. When her clinical course was evaluated retrospectively, it became evident that she had had a similar episode in 2004; at that time, she had developed a mild cough, ground-glass opacities in the lower lobe of the left lung, peripheral eosinophilia, and elevated serum ACE activity ( Fig. 2). All these symptoms remitted spontaneously within several months without treatment.

Once prepared, formulations were stored in tightly closed contain

Once prepared, formulations were stored in tightly closed containers to further reduce the chances of evaporation of ethanol. Optimization of formulations was done by varying the two process parameters i.e. amount of SPC and alcohol. Amount of SPC was varied from 1% to 3% whereas alcohol concentration was varied from

20% to 40%. Different formulations with varying concentrations of SPC and alcohol were characterized for appropriate physicochemical attributes. It is seen from TEM photomicrograph that ethosomal vesicles are unilamellar in nature with near spherical shape (Fig. 2 inset). In contrast to the liposome vesicles that exhibit spherical shapes, ethosomal vesicles are showing some deviation in find more terms of shape, which could be due to absence of cholesterol and presence of ethanol that imparts some level of fluidity to the bilayer membrane. Vesicle size plays an important role in the overall performance of transdermal drug delivery system. Size analysis of the formulations showed that minimum vesicle size was observed to be 120.3±6.1 nm whereas maximum was 410.2±21.8 nm depending on the concentration of SPC and ethanol (Fig. 2). A surface plot was prepared to check the effect of both ethanol concentration and SPC concentration together on vesicle size. A zone of lowest vesicle size having a size range from 100–143 nm is clearly seen in Fig. 3. This particular zone is formed with

formulations having 1–2% of SPC and 35–40% Venetoclax of alcohol. However the lowest size is obtained with 1% of SPC and 40% of alcohol. A zone with the highest size range of 362.5–406.3 nm is formed with 2.5–3% of SPC and 20–25% of alcohol. ON-01910 molecular weight The size of the vesicles

was found to be increased with increasing concentrations of SPC from 1–3% whereas concentration of alcohol affected the vesicle size in the opposite way i.e. higher concentrations of alcohol produced lower vesicle size. It has been reported that high concentration of ethanol confers a surface negative net charge to the vesicular systems by manipulating some surface characteristics, which causes the size of vesicles to decrease [13]. When the amount of SPC was kept at 1% and the concentration of ethanol was increased from 20% to 40%, the size of the vesicles decreased from 212.4±8.3 to 120.3±6.1 nm (Fig. 2). Similarly when the amount of SPC was kept at 2% and 3% and the ethanol concentration was increased in the same manner the vesicle size was decreased from 295.1±14.2 to 147.8±8.3 nm and 410.2±21.8 to 201.2±11.2 nm, respectively (Fig. 2). Encapsulation efficiency is the fraction of total drug entrapped in the ethosomal system. It is an important parameter to evaluate as it determines drug holding capacity and ultimately the delivery potential of dosage form. Zones of different entrapment efficiencies with reference to the concentrations of SPC and ethanol can be identified from the three dimensional surface plot (Fig. 4).

, Paisley, UK) A cell count was then performed using a haemocyto

, Paisley, UK). A cell count was then performed using a haemocytometer, and the cell morphology was examined on cytocentrifuge preparations stained with May Grunwald Giemsa. The purity of PMs was further determined using non-specific esterase staining [16], and the cell viability was determined by trypan blue dye exclusion. At this stage the cell suspension was then kept at 0 °C until assayed. C. albicans (Sigma Chemicals, Poole, UK) was heat killed by placing it in a hot bath at 60 °C for one hour. The yeast particles were then re-suspended and counted before labelling them with fluorescein isothiocyanate (FITC) (Sigma Chemicals, Poole, UK) according to

the method of Ragsdale and Grasso, with some modification [17]. Labelled yeast cells were LDN-193189 research buy LBH589 in vitro re-suspended at 5×107 cells/mL in 20% DMEM-FCS (Life Technologies, Paisley, UK) stored in a light-protected environment at 4 °C. This stock was used within ten weeks

of preparation. Yeast particles opsonisation was performed with pooled human serum (PHS) collected from healthy laboratory staff personnel and stored at −70 °C [18]. The opsonisation step was done one hour before the phagocytosis assay. PMs, which were prepared (1.3.3), were seeded at 0.5–1×106 cells/well in 24-well tissue culture plates (Nunclon, InterMed, Roskilde, Denmark) in RPMI/FCS at 37 °C and 5% CO2 air for two hours to allow for cell adherence. Non-adherent cells were thereafter gently washed out twice with warm HBSS (Life Technologies, Paisley, UK). The labelled

yeast particles were added to wells at a macrophage-to-particles ratio of 1:40 and phagocytosis Mirabegron was allowed to proceed for thirty minutes. In experiments in which GM-CSF was used, the cells were incubated with this cytokine at concentrations of 50, 100 or 500 IU/mL for thirty-six hours, and then phagocytosis was measured [19]. All samples were assayed in duplicates. At the end of this period, the plates were placed on ice and washed twice with ice-cold HBSS, as described previously [17]. Each wash involved gentle pipetting up and down ten times after which the supernatant was discarded. The plates were examined using an inverted microscope between washings to determine the number of internalised particles and to ensure that non-internalised particles, and not cells, were actually washed away. To ensure that there were no free or non-internalised particles left behind, the wells were rinsed once more with the same buffer in some experiments, and the fluorescence of the supernatant was measured using a spectrofluorometer (RF-540 Shimadzu Corp, Kyoto, Japan) at an excitation wavelength of 482 nm and an emission wavelength of 520 nm [17]. Finally cells were lysed using 0.1 N NaOH, and the fluorescence of the liberated intracellular particles in the supernatant was measured using spectrofluorometer (1.3.4).

Participants performed 6 sets of 10 repetitions of


Participants performed 6 sets of 10 repetitions of

maximal isokinetic eccentric knee extension (CSMi Humac NORM, Stoughton, Maryland, USA) at 120° × s−1 with 1 min of rest between each set on each day. This protocol has been used previously to induce muscle damage [13]. Participants were seated and secured to the dynamometer chair as described above but the range of motion of the dynamometer was set between 0° (full knee extension) and 90° (knee flexion) for the eccentric knee extension exercise. The participants were instructed to resist with maximal force against the dynamometer arm as it Ipilimumab manufacturer moved their knee from extension to flexion. The dynamometer motor was used to passively bring the leg back up to the starting position click here after each repetition was completed. All participants were verbally encouraged to exert maximal resistance to the dynamometer throughout each session. A visual analog scale (VAS) was used to assess delayed onset muscle soreness (DOMS). The instrument consists of a 100 mm long line with the words “no pain” on the left hand side and “unbearable pain” on the right hand side of the line. Participants were asked to draw a vertical line on

the scale to rate their current level of pain in their right leg. A pain-rating index was calculated by measuring the distance of the mark from the left hand side of the diagram in millimeters [15]. Pain was assessed before and after each exercise bout as well as 24 h after the final bout with the participants in a standing position. Thigh circumference was measured as an indicator of any edema that may have occurred due to the eccentric exercise intervention. To measure thigh circumference, the halfway distance between the greater trochanter of the femur and the lateral femoral epicondyle on the exercised leg was measured and marked with a horizontal line on each participant using a permanent marker. A measurement of the

circumference of the thigh in centimeters was taken at this horizontal line and recorded in centimeters. The participant remained in a standing position with the thigh muscle relaxed and the feet shoulder width apart while the thigh measurement was taken. Thigh circumference was measured before and after each exercise bout and 24 h after Thymidine kinase the final exercise bout. Range of motion of the knee joint (in degrees) used in the eccentric exercise bouts was assessed with a goniomoter using a protocol previously developed [16]. Briefly, the participant lay prone on the ground with their right leg fully extended. The fulcrum of the goniomoter was placed on the lateral femoral epicondyle of the right leg. The stationary arm of the goniomoter was aligned with the greater trochanter of the femur while the moveable arm was aligned with the lateral fibula. The fully extended knee was considered the starting position of 0°.

Therefore, hypoxia can lead to a rapid increase in HIF-1α protein

Therefore, hypoxia can lead to a rapid increase in HIF-1α protein levels [163], [166], [167] and [168]. Furthermore, HIF-1α up-regulates a number of important factors for tumor expansion, including VEGF, a key factor in tumor angiogenesis [169], [170] and [171]. In several cancers, overexpression of HIF-1α protein has been found to be associated with tumor aggressiveness and with an unfavorable prognosis [159], [172] and [173]. Hypoxia has also been reported to induce wild-type p53 via a

different pathway than DNA-damaging agents [174]. The hypoxic/anoxic induction of p53 selects Olaparib manufacturer for tumor cells that lack functional p53, and hence evidence diminished apoptotic potential [175]. Elevated levels of HIF-1α are noted in various malignant tumors [159], but it is

unclear whether this is so in oral carcinoma. Therefore, we have examined the implications of HIF-1α expression in HOSCC, in vitro and in vivo. NanoCulture plate system was used to duplicate hypoxic condition within tumor mass of living organisms by the three-dimensional cell culture. As the results, we found that HIF-1α regulates the expression of VEGF, and that HIF-1α may be regulated by p53 in SCC of the oral cavity [176] ( Table 5). Although there are numerous anti-cancer drugs, here, we would daringly like to propose the anti-cancer effect of cimetidine, which is an H2R blocker. Because cimetidine is just a stomach medicine, and it is free of side effects Decitabine such as that the other anti-cancer drugs have. Furthermore, H2R blockers are comprised of cimetidine, famotidine, and ranitidine, however, it is reported that only cimetidine has a precise effect as the anti-cancer drug. In addition, although malignant glandular tumors

are known to be generally resistant to radiation therapy and chemotherapy, the efficacy of cimetidine is observed occasionally in the clinical application Reverse transcriptase for glandular tumor. Therefore, if cimetidine has an effect of anti-cancer drug for oral cancer, it will be ideal drug in this lesion. Cimetidine, the oldest histamine type-2 receptor (H2R) antagonist to be used clinically, is commonly prescribed to treat gastro-esophageal reflux disease as well as gastric and duodenal ulcers [177]. Although it is like a half-remembered drug, it has recently been reported that cimetidine improves the survival of patients with malignant tumors [178] and [179], including gastric [180] and colorectal carcinomas [181]. Cimetidine has been shown to inhibit growth of gastrointestinal cancers via several mechanisms including enhancement of immune activity and inhibition of cancer cell proliferation [179]. Therefore cimetidine may act by enhancing the host immune response against tumor cells [182] and [183] or by blocking the cell growth-promoting activity of histamine [183], [184], [185], [186], [187] and [188]. Kobayashi et al.