Tr1 cell clone administration was tolerated and showed dose-dependent efficacy in patients

suffering from severe disease.62 These data represent the first bench-to-bedside test of Tregs as a therapy for IBD and set the stage for more comprehensive trials. Recent work in the field of transplantation and autoimmunity has shown that antigen-specific Tregs are much more effective at preventing graft rejection or diabetes than are polyclonal populations;16 significantly fewer antigen-specific Tregs are required to mediate potent suppression, and the delivery of antigen-specific Forskolin concentration cells decreases the risk of global immunosuppression and the possibility of increased risk of infection and cancer. Notably, antigen-specific Tregs can prevent colitis, as demonstrated by the adoptive transfer of OVA-specific Tregs64 or Tr1 cells,65 but because OVA is unlikely to be Kinase Inhibitor Library high throughput a disease-driving antigen in IBD, the question of whether OVA-specific Tregs would be effective at suppressing established effector responses directed at pathogenic antigens remains outstanding. To develop antigen-specific Treg therapy at least some of the dominant antigens that perpetuate effector T-cell responses in the intestine need to be identified. Using T-cell clones isolated from IBD patients, Duchmann et al.66 found that many of the clones were specific for commensal gut flora, including species of Enterobacteriaceae, Bacteroides

and Bifidobacterium. Corroborating these data, Cong et al.67 found that T cells specific for enteric bacterial flora drive disease in spontaneously colitic C3H/HeJBir mice. It was subsequently demonstrated that

bacterial flagellin, a protein present on all flagellated bacteria including commensal species found in the gut, is a dominant antigen in these mice. In addition, flagellin expressed by a Clostridium species, known as CBir, is targeted by antibodies in colitic mice and humans,68 and transfer of CBir-specific CD4+ T-cell lines into immunodeficient mice causes severe colitis.68 Further evidence that T cells that recognize flagellin are relevant in colitis comes from studies with Escherichia coli-derived flagellin, the delivery of either which exacerbates dextran sodium sulphate-induced colitis in a TLR5-independent manner.69,70 Although this is a new and rapidly evolving field, these data collectively suggest bacterial flagellin as a candidate antigen to target for Treg cellular therapy of IBD. Although dominant antigens that drive IBD are still being discovered, and there are likely to be many different disease-relevant antigens, antigen-directed Treg therapy could currently be tested in a chronic inflammatory gastrointestinal disease that shares similar defects in immune regulation to IBD. Coeliac disease is a chronic immune-mediated inflammatory disorder initiated by wheat gliadin and related proteins in barley and rye.

Strains were evaluated in vitro and in mice. Twenty-two healthy volunteers received single oral doses of either strain in a physiological study of safety, shedding, and immunogenicity. Volunteers were observed in the hospital for seven days and had daily blood cultures, routine safety blood tests (complete blood count with differential; hepatic and renal function), and fecal cultures; none had fever, positive blood cultures, prolonged shedding, or serious or unexpected events. Four of 12 volunteers who received the actA/plcB-deleted strain had minor, transient, asymptomatic serum transaminase elevations (maximum increase 1.4×

upper normal). Six of six volunteers who received ≥4 × 109 colony forming units had detectable mucosal immune responses to listerial antigens, selleck chemicals llc but not to the vectored influenza antigen. Approximately half the volunteers had modest interferon-γ ELISpot responses to a complex listerial antigen, but none had increases over their baseline responses to the influenza antigen. Comparison with prior work suggests that foreign antigen expression, and perhaps also freezing, may adversely affect the organisms’ immunogenicity. Live attenuated bacteria expressing foreign antigens present a promising approach in the development of human immunotherapies and vaccines. Favorable attributes of

Listeria monocytogenes include its innate immunostimulatory Angiogenesis inhibitor properties, efficient cytosolic entry of antigen presenting cells, and its ability to stimulate vigorous CD4 and CD8 T cell responses. L. monocytogenes antigen processing and presentation via

the major histocompatability complex class I pathway makes it an attractive vector for delivery of viral and cancer antigens. Animal studies show that L. monocytogenes vectors can generate T cell-mediated immune responses to lymphocytic choriomeningitis virus, influenza virus, HIV-1 Gag, and human papilloma virus-16 antigens (1–5). Despite these promising results, there are safety concerns related to using recombinant wild type Silibinin L. monocytogenes as a vaccine vector due to the high degree of morbidity and mortality that results from naturally acquired infection. Several groups have developed strategies for attenuating L. monocytogenes by deletion of specific virulence genes (1, 6–8) and two strains have been evaluated in published human studies: ΔactA/plcB (9) and ΔprfA (10) via oral and parenteral routes, respectively. In our prior oral clinical study, 20 healthy adult volunteers received escalating single doses of live attenuated L. monocytogenesΔactA/plcB safely. No individual experienced a serious adverse event. Three of 20 individuals had mild elevations in serum transaminases (maximum 2.5× upper limit of normal) that were temporally associated with vaccination and could not be otherwise explained. Subsequently, another biotechnology group developed an L.

© 2010 Wiley-Liss, Inc. Microsurgery, 2011. “
“Peripheral Enzalutamide nerve surgery performed under unfavorable conditions results

in increased scar formation and suboptimal clinical outcomes. Providing the operated nerve with a protective barrier, reduces fibrosis and adhesion formation and may lead to improved outcomes. The ideal coverage material should prevent scar and adhesion formation, and maintain nerve gliding during motion. Nerve protection using autologous tissues has shown good results, but shortcomings include donor site morbidity and limited availability. Various types of methods and materials have been used to protect nerves. There are both advantages and disadvantages associated with the various materials and techniques. In this report we summarize currently used protective materials applied for nerve coverage under various surgical conditions. © 2013 Wiley Periodicals, Inc. Microsurgery, 2013. “
“Although success of digital replantations

in children has been reported by many authors, the very distal fingertip replantation remains technically demanding. The aim of this article Akt inhibitor is to review our experience with fingertip replantations at or distal to the nail base in pediatric patients and evaluate the clinical outcomes. From October 2000 to May 2007, 12 pediatric fingertips amputated at or distal to the nail base were replanted. Only one artery was anastomosed for revascularization with or without nerve Tolmetin repair; vein drainage was provided by the controlled bleeding technique. Eleven of the 12 replants (91%) survived; one replant of crushed digit failed. An average of 26 month (range, 6 to 36 months) follow-up revealed excellent restoration of finger motion and appearance. The regained static 2-point discrimination (S2PD) sensation was from 3.2 to 5.0 mm (mean, 4.2 mm). Both

the parents and the children were satisfied with the final results. In conclusion, fingertip replantation in children allows good functional and esthetical recovery and should be attempted if technically feasible. © 2010 Wiley-Liss, Inc. Microsurgery 30:380–385, 2010. “
“Severe auricular traumas with extensive involvement of the surrounding structures present with a serious defect necessitating free tissue transfers for reconstruction. In this case report, we present a case of whole left auricle reconstruction with a radial forearm flap prelaminated with porous polyethylene (Medpor®) implant in a 17-year-old female patient. First, a subdermal pouch was fashioned on the volar aspect of the left forearm along the projection of the radial artery and the Medpor implant was placed in this pouch. Four weeks later, the prelaminated radial forearm flap containing the Medpor implant was transferred to the recipient site.

Although ROS is known to cause DNA damage, it also attacks cell

membranes through formation of lipoperoxide (16,17). To explore the possibility that damage to DNA is responsible for the high sensitivity of V. vulnificus to ROS, we examined cytotoxic agents known to be highly specific for DNA, namely, UV, mitomycin C, and methyl methane sulfonate, for their killing effect on V. vulnificus and E. coli. We found that all of these agents kill V. vulnificus much more efficiently than they do E. coli (Fig. 4). Thus, we showed that cells of V. vulnificus are more susceptible to DNA-damaging agents, including Silmitasertib solubility dmso ROS, than are those of E. coli. The primary aim of the present study was to substantiate the remarkable but solitary report of successful HBO treatment of an advanced case of severe V. vulnificus infection (7). Our findings described herein seem to have achieved this purpose. First, we clearly demonstrated the efficacy of HBO treatment alone in a mouse footpad infection model, a beneficial effect being present without chemotherapeutic or surgical

MLN8237 cell line interventions (Fig. 1). In addition, HBO brought about marked viability loss in cells of V. vulnificus in vitro, whereas we did not observe such an effect with E. coli (Fig. 2a). These results support the notion that HBO is an effective therapy for human infections caused by V. vulnificus. However, we wish to emphasize that our observations do not necessarily encourage the use of HBO alone in the treatment of human cases. Rather, HBO therapy should be regarded as a powerful adjunct modality, not only for clostridial gas gangrene (1,2) but also for severe V. vulnificus infection. However, we should be alert for oxygen toxicity, a well-known side effect of HBO, when using HBO against this infection. Two different mechanisms have so far been proposed for the effectiveness of HBO in the treatment of infectious diseases. One is the direct action of oxygen on the offending microbes and the

other is the indirect effect of increased amounts of dissolved oxygen in plasma and infected tissue (1, 2). As to the direct effect of HBO on bacteria, its in vitro bactericidal activity against the obligate anaerobe Clostridium perfringens has been well established (9, 18). Against facultative bacteria, HBO appears to be mostly bacteriostatic as so far studied (19–21), with the exceptions of its bactericidal effect on Vibrio cholerae (comma) (19) and Pseudomonas Calpain aeruginosa (20). On the other hand, the indirect mechanism may include increased microbicidal activity of leucocytes (22) as well as the anti-inflammatory and anti-edemic effects of oxygen, which would help accelerate wound healing (23). One of the interesting aspects of the present study is the mechanism by which the facultative bacterium V. vulnificus behaves as an oxygen-sensitive organism under HBO conditions. Possibly, there is a difference between V. vulnificus and E. coli in oxygen tolerance, which is not manifest in the air, but becomes evident under HBO.

Conventional B-2 cell-derived plasma cells are surface Ig negative, CD19low/negative and express high levels of the plasma cell marker Maraviroc CD138 and slightly higher level of CD43 than B-1 cells (Fig. 5 and 40). BM B-1 cells, >80% of which spontaneously secrete IgM in vitro (Fig. 4C), are surface IgM+IgDlow/negative, and express relatively high levels of CD19 but are CD138− (Figs. 2, 3, 5). Our data are consistent with earlier reports on the phenotype of IgM-secreting B cells 25, 33 and through the use of the allotype chimeras we now identify these BM cells

unequivocally as B-1 cells (Figs. 3 and 4). In addition, as we show here (Figs. 1 and 4), these cells produce antibodies that recognize influenza virus, a specificity we have previously linked to B-1 cell-derived antibodies 5, 26, 27. Staining with antibodies recognizing a B-1a cell-specific Ig-idiotype (T-15) binding to phosphorylcholine, as well as staining with phosphatidylcholine-containing liposomes identified small numbers of BM B-1 cells (data not shown), further confirming their similarity to known B-1 cells with regard to specificity. Notably, these CHIR-99021 nmr cells are distinct from the IgMloIgDhi

sinusoidal BM B cells, which were described recently as rapid IgM secreters following challenge with blood-borne T-independent antigens 42. FACS-sorting experiments did Forskolin cell line not reveal significant spontaneous IgM secretion among IgMloIgDhi B cells in our

non-challenged mice (Fig. 2). In contrast to BM and spleen, PerC B-1 cells from BALB/c mice or from allotype chimeras were not significant sources of spontaneous IgM secretion (Figs. 1 and 4). Thus, our data are consistent with several in vivo studies that indicated the inability of PerC B-1 cells to produce natural IgM 33, 36, 37. It is remarkable, however, that PerC B-1 cells secrete or shed small amounts of IgM, resulting in large numbers of pinhead-size ELISPOTs (Fig. 1), also noted by others 31, 32, without significant amounts of secreted product amassing in the culture supernatants (Figs. 1 and 3). Such “leakiness” of B cells was not noted for cells harvested from any other tissue, for example the PLNs (Fig. 1). This might explain the apparent discrepancies in the literature regarding IgM secretion by PerC B-1 cells 31–37. Counting of these very small dots by PerC B-1 cells, might lead to an over-estimation of the ability of these cells to secrete significant amounts of natural IgM.

A schematic representation of “Injury ABC294640 concentration types and reconstruction algorithm” is shown in Figure 2. Experience on intraoperative vascular pedicle damage during axillary lymph-node dissection by general surgeon is reported and an algorithmic approach regarding types of injuries and available options to repair them in attempt to salvage the flap is developed. The knowledge of what to expect and what to do,

may reduce the incidence of flap loss and reconstruction failure, thus saving the patient from the additional stress of a second procedure. Every surgeon must be aware of such complications and of the available surgical strategies, being then adequately skilled in the different techniques of breast reconstruction including Decitabine supplier microvascular surgery which was required to re-establish blood flow in our cases. “
“The transjugular portosystemic shunt, widely used to treat portal hypertension today, may increase the risk of encephalopathy

and reduce effective hepatic flow. To address these issues, a strategy to produce a portocaval shunt (PCS) with hepatic function using intestinal grafts was conceived, and rat models were developed. We transplanted ileal grafts from wild-type and luciferase transgenic Lewis rats to wild-type Lewis rats, anastomosing the graft mesenteric artery (SMA) and portal vein (PV) to the recipient PV trunk and inferior vena cava, respectively. Recipient survival was significantly longer in the partial PCS model, ADAMTS5 in which the graft SMA was anastomosed to the recipient PV trunk in an end-to-side fashion, than in the total PCS model, with the end-to-end anastomosis. In the partial PCS model, histological and luminescence analyses showed graft survival for 1 month. These results suggest that intestinal grafts can be maintained in the particular conditions required for our strategy. © 2010 Wiley-Liss, Inc. Microsurgery,

2010. “
“The aim of this study was to evaluate long-term regenerative capacity over a 15-mm nerve gap of an autologous nerve conduit, the biogenic conduit (BC), 16 weeks after sciatic nerve transection in the rat. A 19-mm long polyvinyl chloride (PVC) tube was implanted parallely to the sciatic nerve. After implantation, a connective tissue cover developed around the PVC-tube, the so-called BC. After removal of the PVC-tube the BCs filled with fibrin (n = 8) were compared to autologous nerve grafts (n = 8). Sciatic functional index (SFI) was evaluated every 4 weeks, histological evaluation was performed at 16 weeks postimplantation. Regenerating axons were visualized by retrograde labelling. SFI revealed no significant differences.

Therefore hypertension usually precedes the onset of microalbuminuria.3 BP control modulates selleck the progression not only of microangiopathy (diabetic kidney disease and retinopathy) but also of macroangiopathy (Coronary heart disease (CHD) and

stroke). In microalbuminuric people with type 2 diabetes, observational studies have shown an association between poor glycaemic control and progression of albuminuria. A number of studies have identified a strong independent association between hyperglycaemia and the rate of development of microvascular complications.4 The large observational WESDR study5 indicated an exponential relationship between worsening glycaemic control and the incidence of nephropathy as well as retinopathy and neuropathy. The UKPDS has clearly shown the importance of targeting glycosylated haemoglobin (HbA1c) levels close to normal (HbA1c < 7.0%) in people with type 2 diabetes. A modest decrease in HbA1c over 10 years from 7.9 to 7.0% lowered the risk of microvascular endpoints

with the onset of microalbuminuria being reduced by 25%.6 These findings are supported by a study of intensified glycaemic control in non-obese Japanese click here subjects with type 2 diabetes.7 In the UKPDS, there was no significant reduction in the risk of progression from microalbuminuria to proteinuria with intensive blood glucose control.8 The AusDiab study collected information on albuminuria, measured as a spot albumin: creatinine ratio (ACR) (mg/mmol) with microalbuminuria being between 3.4 and 34 mg/mmol and macroalbuminuria at >34 mg/mol.9 The prevalence of albuminuria increased with increasing glycaemia. People with diabetes and impaired glucose tolerance had an increased risk for albuminuria compared with those with normal glucose tolerance, independent of other known risk factors for albuminuria (including age and sex). Hyperglycaemia is an important determinant of the progression of normoalbuminuria to microalbuminuria in diabetes.

Protirelin Strict blood glucose control has been shown to delay the progression from normoalbuminuria to microalbuminuria or overt kidney disease6 and from normo- or microalbuminuria to overt kidney disease.7 The influence of intensive glycaemic control is greatest in the early stages of CKD although some observational studies suggest an association of glycaemic control with the rate of progression of overt kidney disease and even end-stage kidney disease (ESKD).10 The American Heart Association (AHA) has undertaken a review of the DCCT, UKPDS, ACCORD, ADVANCE and VA Diabetes trials and on the basis of the review issued a Scientific Statement addressing intensive glycaemic control in relation to cardiovascular events.11 While the AHA review is focused on cardiovascular events, the statement is relevant to the consideration of the management of CKD given the strong association between CKD and CVD in people with type 2 diabetes.

In addition, we observed that the PKC activator, PMA, as well as a bacterial fermentation end product, butyrate, also regulated TSLP expression both at the mRNA and protein level. Moreover, a strong synergistic effect between PMA and butyrate

was observed. The latter effect may be physiologically relevant given the major biological function of butyrate as an energy source in the colon [30] as well as its function as an epigenetic regulator [31]. As expected, stimulation of IECs by IL-1 induced NF-κB translocation into the nucleus and TSLP transcription involving IKK-β activity as revealed by the specific inhibition induced by Bay 11–7082. Clearly, the functional importance of both p38 and PKA was also identified using SB203580 and H-89, respectively. Conversely, extracellular signal-regulated kinase (ERK) PI3K inhibitor had little effect since UO126 barely inhibited TSLP transcription. We first postulated that both p38 and PKA may act independently of IKK-β involvement since their specific pathway inhibitors were effective in the presence of Bay 11–7082, whereas UO126 had no effect. However, when transient transfections were performed with a 4 kb TSLP-promoter region, mutated for NF2 binding site, the stimulatory effect of IL-1 was completely abolished;

thus Cell Cycle inhibitor arguing for a NF-κB only dependent regulation. We present in Figure 7 our working hypothesis that can explain the overall results obtained in IL-1-dependent TSLP regulation. Considering that, in the presence of BAY 11–7082, the effects of both p38 and PKA inhibitors are still apparent, we can argue that, since BAY 11–7082 has an IC50 of 10 μM [32], at the concentration 20 μM used Tenofovir cell line in the current study, IKK-β may only be partly inhibited and that the remaining TSLP transcription activity is still mediated by IKK-β. This has been verified using a NF-κB-dependent SEAP reporter system [33]. In fact, at the 20 μM concentration, BAY 11–7082 only inhibited IL-1-dependent NF-κB activation

by about 60% in Caco-2 cells. To explain the effects of the p38 inhibitors, our hypothesis is that IL-1 is activating IKK-β by two separate modes; first via the classical IL-1 receptor associated kinase/TGF-β activated kinase (IRAK/TAK) dependent pathway and second via a MKK/p38-dependent pathway as revealed previously for IL-6 [34]. Thus, inhibition of p38 resulted in a decreased TSLP expression due to a reduced activation of IKK-β, and enhances BAY 11–7082 direct inhibitory activity. Considering the involvement of PKA, it has been shown that PKA can also interfere with the NF-κB pathway; indeed PKA was revealed to phosphorylate p65 in a cAMP-independent manner therefore increasing transcriptional activity [35]. Our results argue for a similar regulation of TSLP transcription in human IECs. Recently, TSLP has been shown to be regulated by NF-κB in both human and mice airway epithelial cells [16]. A site located at –3.

As seen in Figure 1, five RCTs involving 263 patients reported Ccr, the meta-analysis showed that the Ccr level was significantly higher in the groups receiving calcium disodium EDTA as compared with the groups receiving placebo (SMD,

0.68; 95% CI, 0.43 to 0.93; P < 0.00001). These results suggest that calcium disodium EDTA chelation therapy has a significant benefit of retarding the progression SB203580 manufacturer of chronic kidney disease in patients with measurable body lead burdens. However, the pooled estimate showed that the calcium disodium EDTA treatment group did not have a significant decrease in urine protein level compared with that of the control group (SMD, −0.17; 95% CI, −0.46 to 0.11; P = 0.24, Fig. 1). The first reported case of nephrotoxicity associated with lead was described more than a hundred years ago. From then on, exposure to lead has been thought of as a risk factor for kidney injury. Because blood lead levels indicate only recent exposure to lead, LDE225 mw the level of body lead burden is considered as a more accurate indicator

reflecting the lead load in the human body, and urinary lead excretion <600 μg/72 h after calcium disodium EDTA chelation therapy is considered as a normal body lead burden. However, it was found that lead has a direct toxic effect on the kidney even at ‘normal or acceptable’ levels.[2, 3] The pathogenesis of nephrotoxic effects of lead is mainly related to direct toxicity, inflammation and oxidative stress.[2,

13, 14] A growing body of research has shown that lead exposure is a reversible risk factor for CKD progression,[4-9] nonetheless, the optimal strategy to treat lead nephrotoxicity remains uncertain. Chelating agents such as calcium disodium EDTA are widely used to remove toxic metals, and this therapy could exert long-term antioxidant, anti-inflammatory effects.[15] However, lead chelation is controversial owing to the potential of its use in lieu of exposure reduction. In addition, cases of acute tubular necrosis have been reported following early clinical use of calcium disodium EDTA that involved very large doses.[16] Fortunately, adverse renal effects have not been observed Bay 11-7085 at low levels of exposure such as in the trials included in our meta-analysis. The main finding of the current meta-analysis is that calcium disodium EDTA chelation therapy could effectively delay the progression of chronic kidney disease among patients with measurable body lead burdens by increasing the levels of GFR and Ccr. There is no conclusive evidence that chelation therapy with calcium disodium EDTA reduces proteinuria. However, our findings indicate the need to be confirmed by more larger randomized clinical trials. There are several important potential study limitations to this meta-analysis. First, most of the included studies were small-scale studies that may have had patient selection and treatment bias. Second, most studies were not blinded.

Orf2 was proposed to be formyltransferase for synthesis of dTDP-d-Qui3NFo from dTDP-d-Qui3N. Therefore, we suggested that orf3, orf4, orf5, and orf2 are involved in the synthesis of dTDP-d-Qui3NFo and named them rmlA, qdtA, qdtB, and qdtF, respectively. Orf11 shares 76% GSK3235025 manufacturer identity or 89% similarity to UDP-glucose 6-dehydrogenase (Ugd) of Edwardsiella ictaluri, which is responsible for the synthesis of UDP-d-GlcA from UDP-d-Glc (Stevenson et al., 1996). Therefore, orf11 was proposed to be responsible for the synthesis of UDP-d-GlcA and named ugd. Both Orf13 and Orf14 belong to the NAD-dependent epimerase/dehydratase family (Pfam01370, E value = 3× e−23

and 3 × e−45, respectively). Orf13 shares 78% identity to UDP-N-acetylglucosamine 4-epimerase (Gne) of P. mirabilis. In Providencia (Ovchinnikova et al., 2012), as in most other Enterobacteriaceae members studied (Valvano, 2011), the O-unit synthesis is likely initiated Gemcitabine research buy by transfer of GlcNAc-1-phosphate or GalNAc-1-phosphate to the undecaprenol phosphate (UndP) lipid acceptor. Recent

biochemical studies showed that Gne from E. coli O157 is capable of interconverting GlcNAc-P-P-Und and GalNAc-P-P-Und rather than functions as a UDP-GlcNAc/UDP-GalNAc epimerase (Rush et al., 2010). As GalNAc is evidently the first monosaccharide of the P. alcalifaciens O40 O-unit (Ovchinnikova et al., 2012), it is not excluded that Orf13 is responsible for the synthesis of GalNAc-P-P-Und from GlcNAc-P-P-Und too. The fourth sugar component Dapagliflozin of the O-unit is d-galactose. Seventy-four percent identity was observed for Orf14 compared to UDP-galactose 4-epimerase (GalE)

of P. mirabilis. Therefore, orf14 was named galE. However, it should be noted that in most other Providencia strains studied, galE is located at the 3′ end of O-antigen gene clusters between cpxA and yibK independently of the presence of galactose in the O-unit (Ovchinnikova et al., 2012). Orf10 shares 28% identity or 48% similarity to a putative galactoside acetyltransferase of Bacteroides thetaiotaomicron, and the corresponding gene was named wpaC. The presence of this gene is consistent with partial O-acetylation of the O-unit; however, the position of O-acetyl group on the Gal residue was not confirmed chemically. The transfer of a 2-acetamido sugar 1-phosphate to UndP is mediated by WecA, which also takes part in the enterobacterial common antigen (ECA) synthetic pathway. The wecA gene encoding this enzyme is located in the ECA biosynthesis gene cluster (Alexander & Valvano, 1994). Therefore, three individual glycosyltransferases were expected to assemble the UndPP-linked tetrasaccharide O-unit of P. alcalifaciens O40. Both Orf7 and Orf12 belong to the glycosyltranferase group 2 family (Pfam00535, E value = 2 × e−16 and 9 × e−33, respectively).