Methylene blue - an emerging vasopressor

Abstract

Methylene blue (MB) is being investigated as a useful adjunct in the management of vasoplegic and septic shock due to its targeted inhibition of the nitric oxide-soluble guanylate cyclase pathway. Early studies demonstrated improved systemic vascular resistance and reduced catecholamine requirements, laying the foundation for later randomized trials. The pivotal Ibarra-Estrada trial (2023) confirmed that early MB infusion decreases time to vasopressor discontinuation and increases vasopressor-free days. Subsequent data further support MB’s hemodynamic benefits in refractory distributive shock. Dosing regimens need to be refined, with cumulative doses of 2-4 mg/kg demonstrating optimal safety and efficacy currently. MB retains a favorable safety profile with minimal adverse effects when used within recommended limits. Additional applications include cardiac surgery vasoplegia, anaphylaxis, calcium channel blocker toxicity, ifosfamide encephalopathy, and reperfusion vasoplegia in liver transplantation. Overall, current evidence supports MB as a rational, mechanism-based adjunctive vasopressor in carefully selected patients with vasodilatory shock however a large randomised controlled trial and further evidence is needed to answer many lingering questions.

Key Words: Methylene blue; Vasodilatory shock; Septic shock; Nitric oxide pathway; Vasoplegia; Adjunctive vasopressor therapy; Ifosamide

Core Tip: Methylene blue (MB) is a mechanistically rational adjunct vasopressor that targets nitric oxide (NO)-mediated vasodilation in septic shock. When used early and within safe cumulative dose (≤ 4 mg/kg), it consistently reduces vasopressor requirements without toxicity. Unlike non-selective NO synthase inhibitors, MB selectively modulates the NO-guanylate cyclase-cyclic guanosine monophosphate pathway by inhibiting only the inducible NO synthase isoform of NO synthase, preserving physiological constitutive functions. Current evidence supports its use as an adjunct rather than rescue therapy, pending results from ongoing randomized trials. In this narrative review we present the facts and data on MB with no fluff!

INTRODUCTION

Methylene blue (MB) (methylthioninium chloride) (C₁₆H₁₈ClN₃S) was discovered by Heinrich Caro in 19^th century, a German scientist. Its use in septic shock has seen a tottering course till the Ibarra-Estrada trial in 2023 leading to an increasing number of publications, interest and use of MB as a vasopressor.

MECHANISM OF ACTION OF MB IN SEPTIC SHOCK

Septic shock is characterized by profound vasodilatation predominantly driven by excess nitric oxide (NO) production, with approximately 30% of patients also exhibiting concomitant myocardial depression. This pathological vasoplegia is primarily mediated by inducible NO synthase (iNOS), which is markedly upregulated during systemic inflammation.

NO is synthesized by NO synthase (NOS), which exists in two major functional forms: Constitutive NOS (cNOS) and iNOS. cNOS comprises endothelial NOS and neuronal NOS and is responsible for physiological vascular homeostasis, endothelial integrity, basal blood pressure regulation, and neuronal signaling[1-3]. These pathways are essential for normal cardiovascular and cellular function and remain active under basal conditions.

In contrast, iNOS is minimally expressed in healthy states but is robustly induced during sepsis by inflammatory cytokines, endotoxins such as lipopolysaccharide, and other mediators. Once activated, iNOS generates large and sustained amounts of NO, leading to pathological vasodilation, impaired vascular responsiveness, and myocardial depression. Importantly, in septic shock, it is the excess NO derived from iNOS - rather than cNOS - that drives hemodynamic collapse, while cNOS pathways remain essential and largely unchanged for physiological homeostasis.

This distinction has critical clinical relevance. Prior attempts at non-selective NOS inhibition, most notably with N(G)-methyl-L-arginine hydrochloride (L-NMMA) (546C88), suppressed both iNOS and protective cNOS signaling and were associated with increased harm and mortality, leading to early termination of clinical trials[4]. These findings underscored the dangers of global NOS blockade and halted further development of non-selective strategies.

MB differs mechanistically from non-selective NOS inhibitors. Rather than globally suppressing NOS activity, MB preferentially attenuates iNOS-mediated NO signalling and its downstream effects, while preserving cNOS function. This selective modulation provides a mechanistic rationale distinct from prior failed approaches.

Downstream, NO activates guanylate cyclase, resulting in increased cyclic guanosine monophosphate (cGMP) production and subsequent relaxation of vascular smooth muscle cells and cardiomyocytes. MB inhibits guanylate cyclase and reduces cGMP generation; however, this effect is likely secondary and of lesser importance compared with its upstream attenuation of cytokine-driven iNOS activation and excessive NO production[5,6]. The differential effects of MB on the NO-guanylate cyclase-cGMP pathway, contrasted with non-selective NOS inhibition, are illustrated in Figures 1 and 2 which contrast selective inhibition of the iNOS-NO-cGMP pathway by MB with non-selective NOS inhibition by L-NMMA, which explains the divergent clinical outcomes observed and the need to understand the use of MB.

Figure 1 Mechanism of methylene blue in selectively inhibiting inducible nitric oxide synthase that prevents nitric oxide generation. It does not block production of other isoform i.e. constitutive nitric oxide synthase which is required for various constitutive functions in the body. Further methylene blue also inhibits the activation of cyclic guanosine monophosphate by guanylate cyclase, ultimately all this prevents the extreme vasodilatation as shown in Figure 2. MB: Methylene blue; NO: Nitric oxide; iNOS: Inducible nitric oxide synthase; cGMP: Cyclic guanosine monophosphate.

Figure 2 Non selective inhibitor N(G)-methyl-L-arginine hydrochloride (molecule 546c88) inhibiting nitric oxide synthase and thus all downstream nitric oxide synthase isoforms resulting in complete blockade of production of nitric oxide causing increased mortality López et al[4]. NOS: Nitric oxide synthase; iNOS: Inducible nitric oxide synthase; cNOS: Constitutive nitric oxide synthase; eNOS: Endothelial nitric oxide synthase; nNOS: Neuronal nitric oxide synthase.

Hemodynamic effects of MB

Beyond its effects on vascular tone, MB has been shown to exert direct effects on cardiac function and the pulmonary circulation. Excess NO is a well-recognized myocardial depressant and pulmonary vasodilator. In septic shock, inhibition of pathological NO excess by MB counteracts these effects, leading to improved left and right ventricular contractility and an increase in pulmonary vascular resistance.

These changes enhance ventriculo-arterial coupling and are associated with improvements in key hemodynamic indices, including cardiac index, left and right ventricular stroke work indices, mean pulmonary artery pressure, and pulmonary vascular resistance index. Such effects were consistently demonstrated in two well-conducted invasive hemodynamic studies by Kirov et al[7] and Donati et al[8], providing mechanistic support for the observed catecholamine-sparing effects of MB in septic shock.

Early experience with MB: Case reports and initial trials

The initial clinical use of MB as a vasopressor occurred in patients with vasoplegia following cardiopulmonary bypass during cardiac surgery. Although the underlying trigger differed, the pathophysiological mechanism was analogous to septic shock - excess NO production driven by cytokine release associated with surgical trauma and exposure to the bypass circuit rather than infection[9]. In this setting, a single bolus of MB was often sufficient to reverse vasoplegia, as postoperative vasoplegia typically resolves within 6 hours[10,11].

The first reported use of MB in septic shock was by Schneider in 1992, who successfully weaned vasopressors within 12 hours in two patients[12]. Subsequently, Brown et al[13] reported prolonged MB infusion for up to 44 hours with successful norepinephrine tapering in septic shock. These reports were followed by several case series and small observational studies suggesting hemodynamic benefit.

Two small randomized controlled trials by Kirov et al[7] and Memis et al[14] demonstrated reductions in vasopressor requirements with MB use. These remained the only randomized studies evaluating MB itself until the late 2000s. In contrast, a large multicenter randomized trial conducted by López et al[4] enrolled 797 patients with septic shock and followed earlier phase II studies[15,16]. This trial evaluated the non-selective NOS inhibitor L-NMMA (molecule 546C88), not MB. The study was terminated early because of increased 28-day mortality in the treatment arm (59% vs 49%, P < 0.001)[4]. Although no definitive mechanism was identified, the authors postulated that excessive vasoconstriction [target mean arterial pressure (MAP) 70-90 mmHg] and myocardial dysfunction may have contributed, as the excess mortality was predominantly cardiovascular.

Critically, L-NMMA inhibits both inducible and cNOS isoforms, suppressing protective cNOS signaling alongside pathological iNOS activity. In contrast, MB selectively attenuates iNOS-mediated NO signaling while sparing cNOS pathways essential for vascular and cellular homeostasis. Global inhibition of NO is known to have deleterious multisystem effects, and it is therefore plausible that the excess mortality observed in the López trial was related to non-selective NOS inhibition rather than NO modulation per se.

Observational data and post-2007 evidence

A large retrospective cohort study published in 2022 evaluated 223 patients who received MB for refractory shock, the majority of whom had septic shock[17]. The median bolus dose was 1.7 mg/kg, with a continuous infusion rate of 0.45 mg/kg/hour. Hemodynamic responders - defined by a MAP increase > 10% - demonstrated a reduction in norepinephrine requirements within one hour of MB administration. Although overall mortality was not significantly different, responders showed a trend toward improved outcomes.

Notably, among responders receiving continuous renal replacement therapy (CRRT), intensive care units mortality was lower. The authors hypothesized that reduced clearance of MB during CRRT may lead to higher effective plasma concentrations. Supporting this observation, another study demonstrated a potential renoprotective effect of MB, with improved creatinine clearance and reduced urinary markers of tubular injury, which may partially explain improved outcomes in responders[18].

Overall, early case reports and small trials consistently demonstrated rapid catecholamine sparing with MB use; however, conclusions were limited by small sample sizes, heterogeneous dosing strategies, and variability in timing of administration.

The pivotal 2023 Ibarra-Estrada trial

The randomized trial by Ibarra-Estrada et al[19] in 2023 remains the most robust evidence supporting MB as an adjunct vasopressor in septic shock. This placebo-controlled randomized study enrolled 91 patients with early septic shock, with MB initiated within the first 24 hours. The trial was not powered to detect mortality differences.

All patients were receiving norepinephrine at enrollment. MB was administered as 100 mg infused over 6 hours daily for three consecutive days, resulting in a cumulative exposure of approximately 3.6 mg/kg. Vasopressin was added when norepinephrine requirements reached 0.25 μg/kg/minute.

MB significantly reduced vasopressor duration, shortening time to vasopressor discontinuation by nearly 25 hours. Vasopressor-free days at day 28 increased, and intensive care units and hospital length of stay were modestly reduced. There was no effect on ventilator duration or mortality, and importantly, no MB-related adverse events were observed. This trial re-established MB as a structured early adjunct rather than a rescue therapy and revived global interest in its use.

Developments after the Ibarra-Estrada trial: Expanding but limited evidence

Following publication of the Ibarra-Estrada trial, few additional randomized studies have emerged. Post-2023 literature is dominated by pharmacologic reviews, meta-analyses, and small observational datasets. A consistent pattern is evident: MB is hemodynamically effective, appears safe, and is most beneficial when administered early.

Shaker et al[20] conducted a randomized trial in a cancer patient cohort with shock - predominantly septic - in which both low- and high-dose MB regimens were evaluated, demonstrating efficacy and safety. Multiple meta-analyses by Ballarin et al[21], Fernando et al[22], Huang et al[23], and Zhao et al[24], encompassing pooled patient numbers exceeding 1000, reached similar conclusions. MB reliably increases MAP, shortens vasopressor duration, and may confer a modest mortality benefit when used as a second-line adjunct rather than late rescue therapy. However, these analyses uniformly noted significant heterogeneity and the predominance of small studies, limiting the strength of conclusions.

Importantly, across studies evaluating MB itself, serious adverse effects remain rare (Tables 1 and 2). The López et al[4] trial remains a notable outlier, but it evaluated non-selective NOS inhibition rather than MB and should not be extrapolated to MB therapy.

Table 1 A summary of trials from 2023 to 2025.

Ref.	Study design/population	Methylene blue regimen	Comparator	Major findings/clinical outcomes	Safety/adverse events	Key interpretation
Ibarra-Estrada et al[19], 2023	Randomised controlled trial (n = 91); adult septic shock (sepsis-3)	100 mg IV bolus infusion over 6 hour daily × 3 days (approximately 3.6 mg/kg total)	Placebo (saline)	Decreasing time to vasopressor discontinuation (69 hours vs 94 hours, P < 0.001); increasing vasopressor-free days; intensive care units and hospital LOS decrease; no mortality difference	No significant toxicity; no methemoglobinemia	Early adjunctive MB within 24 hours shortened vasopressor duration without added risk
Fernando et al[22], 2024	Systematic review + meta-analysis (> 1000 patients)	Pooled 1-4 mg/kg bolus ± 0.25-1 mg/kg/hour infusion	Standard therapy	Decreasing vasopressor duration (approximately 20 hours); trend 28-day mortality decrease (RR: 0.82); mean arterial pressure increase	No increase in adverse events	MB benefit most pronounced when used early as adjunct
Ballarin et al[21], 2024	Systematic review + meta-analysis (15 studies/1200 patients)	1-4 mg/kg bolus ± continuous infusion	Placebo/standard care	Decreasing norepinephrine dose (-16%); increasing urine output and CrCl; no mortality harm	No toxicity reported	Confirms hemodynamic benefit and renal protection
Arias-Ortiz and Vincent[25], 2024	Narrative review/mechanistic update	Summative PK-PD review; discussed dose-response	-	Defined biphasic response; supports 2-4 mg/kg as effective and safe	Warns against > 7 mg/kg due to splanchnic vasoconstriction	Clarifies pharmacologic rationale and dose limits
Shaker et al[20], 2025	Double-blind randomised controlled trial (n = 90) - cancer patients with septic shock	Low dose: 1 mg/kg bolus + 0.25 mg/kg/hour × 72 hour. High dose: 4 mg/kg bolus + 0.25 mg/kg/hour × 72 hours	Placebo (saline)	Both MB arms decrease vasopressor duration and increasing vasopressor-free days vs placebo (P < 0.05); trend mortality decrease (P = 0.083)	No methemoglobinemia or hemodynamic instability	Confirms efficacy up to 4 mg/kg; possible dose ceiling effect

IV: Intravenous; LOS: Length of stay; MB: Methylene blue; RR: Relative risk; PK-PD: Pharmacokinetics-pharmacodynamics.

Table 2 Adverse effects[28-31].

Adverse effect	Comments/mechanism
Hemolysis (glucose-6-phosphate dehydrogenase deficiency)	Contraindicated; oxidative stress triggers hemolysis
Methemoglobinemia	< 2% incidence; dose-related
Serotonin syndrome	Occurs with MAOI/SSRI co-administration
Green-blue discoloration	Urine/secretions/skin; due to metabolites; affects pulse oximetry
Pulse oximetry artifacts	Spurious low SpO2; due to light absorption
Fall in P/F ratio	Possibly from pulmonary vasoconstriction → V/Q mismatch. Rare-high doses 31
Splanchnic vasoconstriction	At doses > 7 mg/kg (possibly > 4 mg/kg)
Extravasation injury	Can cause necrosis; central line preferred
ECG changes, diaphoresis, dyspnea	Reported at higher doses
Pregnancy risk	Avoid use; insufficient safety data

MAOI/SSRI: Monoamine oxidase inhibitor/selective serotonin reuptake inhibitor; P/F: PaO₂/FiO₂ ratio; V/Q: Ventilation/perfusion; ECG: Electrocardiogram.

Dosing insights

Recent pharmacologic analyses highlight why dosing and cumulative exposure are critical determinants of MB efficacy and safety. At lower doses, MB selectively modulates the NO-soluble guanylate cyclase-cGMP pathway, restoring vascular tone while preserving regional and microvascular perfusion. In contrast, higher cumulative doses may lead to excessive vasoconstriction, particularly in the splanchnic circulation, and increase the risk of dose-related toxicities such as methemoglobinemia[25].

Current practical dosing strategies fall into two broad categories. The most evidence-based regimen is derived from the Ibarra-Estrada trial, in which MB was administered as 100 mg infused over 6 hours daily for three consecutive days, resulting in a cumulative exposure of approximately 3.6 mg/kg. Alternative regimens reported in earlier studies include a bolus dose of 1-4 mg/kg, with or without a subsequent infusion for 12-24 hours[20]. Both approaches achieve comparable increases in MAP, provided cumulative exposure remains moderate.

At present, available data suggest that cumulative doses exceeding 7 mg/kg may be associated with adverse effects, while robust safety data beyond this threshold are lacking. Consequently, the optimal dosing strategy and the precise upper safe limit remain unsettled. The maximum duration of MB administration reported in the literature to date is approximately 120 hours[26].

Pharmacokinetics

MB follows a two-compartment pharmacokinetic model, with a distribution half-life of approximately 5-6 hours and a terminal elimination half-life of around 20 hours. Hemodynamic effects typically begin within the first hour of administration but are not immediate. MB undergoes hepatic reduction to leucomethylene blue, producing the characteristic blue-green discoloration of urine and secretions. Approximately 75% of the drug is excreted as metabolites and 25% unchanged[27].

Patients receiving CRRT may achieve higher effective MB concentrations, potentially explaining the enhanced hemodynamic responses observed in earlier observational cohorts. As noted by Naoum et al[17], “observed enhanced responses in CRRT patients may relate to altered volume of distribution or clearance; definitive pharmacokinetic data are lacking”. Dosing in hepatic dysfunction remains uncertain, given hepatic metabolism of the drug, as does the maximum safe duration beyond the approximately 120-hour exposure currently reported[28].

Microcirculatory effects?

Across studies, MB consistently increases systemic vascular resistance, improves mean arterial pressure, preserves cardiac index, and reduces norepinephrine requirements[7,8,22,23,25]. Ballarin et al[21] reported an approximately 16% reduction in norepinephrine-equivalent dose within 6 hours of MB administration. Improvements in urine output and creatinine clearance have also been described in several cohorts.

However, improvements in macro-hemodynamic parameters do not necessarily translate into proven microcirculatory benefit. Whether MB meaningfully improves oxygen delivery or tissue oxygen utilization remains uncertain[25]. Modest reductions in serum lactate reported in some studies, including Zhao et al[24], are insufficient to establish a definitive microcirculatory effect because of small sample sizes and substantial heterogeneity.

Safety profile (Table 2)

MB remains a generally safe drug when used within recommended dosing ranges. Hemolysis in patients with glucose-6-phosphate dehydrogenase deficiency and serotonin syndrome with concomitant monoamine oxidase inhibitors or selective serotonin reuptake inhibitors represent the most clinically significant risks. Methemoglobinemia is uncommon and typically mild. Blue-green discoloration of urine, secretions, and skin is frequent and may interfere with pulse oximetry readings[28-31].

At cumulative doses above 7 mg/kg - and possibly above 4 mg/kg in susceptible individuals - splanchnic vasoconstriction and vasopressor-like toxicities may occur[32]. Tissue necrosis from extravasation underscores the importance of central venous administration.

OTHER CLINICAL USES

Anaphylactic shock

MB has been successfully used in refractory anaphylactic shock, including reactions related to protamine, aprotinin, contrast agents, and antibiotics. In several reports, patients unresponsive to fluids, vasopressors, antihistamines, and corticosteroids demonstrated rapid hemodynamic improvement following MB bolus administration[33,34]. Histamine upregulates endothelial NOS expression, increasing NO production; inhibition of guanylate cyclase attenuates histamine-mediated vasodilation[35-38]. Del Duca et al[34] described two cardiac surgery patients with severe anaphylaxis in whom a single 100-mg MB bolus rapidly restored blood pressure and resolved urticaria.

Liver transplantation

MB has demonstrated benefit in vasoplegia associated with orthotopic liver transplantation, particularly during the reperfusion phase. Longhitano et al[39] conducted the only randomized controlled trial in this setting, administering 1.5 mg/kg MB before reperfusion in 36 patients, resulting in attenuation of reperfusion-related hypotension, increased systemic vascular resistance, lower lactate levels, and reduced epinephrine requirements. More recently Harding et al[40] reported further support for its utility in refractory post-reperfusion vasoplegia.

Refractory drug poisoning

MB has been reported as adjunctive therapy in refractory shock due to calcium channel blocker overdose, particularly amlodipine toxicity[41], with occasional reports of benefit in metformin-associated shock.

Ifosfamide-induced encephalopathy

MB has also been used in ifosfamide-induced encephalopathy, a toxic metabolic encephalopathy mediated by chloroacetaldehyde-induced mitochondrial dysfunction and oxidative stress. MB acts as an alternative electron acceptor, inhibits formation and effects of chloroacetaldehyde, and improves mitochondrial function. A review by Abahssain et al[42] identified 58 reported cases, with 65.5% receiving MB and 75.6% demonstrating clinical improvement, particularly when administered early. Robust prospective data, however, remain lacking.

KEY MESSAGE

MB inhibits pathological NO signaling primarily by attenuating cytokine-driven iNOS activation and, to a lesser extent, by blocking guanylate cyclase-mediated cGMP generation, thereby reducing vasodilation and myocardial depression. Over two decades of published data support MB as an adjunct vasopressor in vasodilatory shock - including septic, cardioplegic, and anaphylactic shock - without major adverse effects when used within recommended dosing ranges. Despite growing evidence, MB has not been incorporated into guidelines because of uncertainties regarding optimal dosing, duration, and its impact on oxygen delivery and utilization. Current evidence supports MB as an adjunct vasopressor in septic shock at doses of 100 mg infused over 6 hours daily for three days or a 2 mg/kg bolus followed by infusion (0.25-1 mg/kg/hour), with further clarification needed regarding standardized protocols. MB causes blue-green discoloration of secretions and skin and may interfere with pulse oximetry; higher doses increase the risk of methemoglobinemia, hemolysis (especially in glucose-6-phosphate dehydrogenase deficiency), and mesenteric ischemia. Ongoing and future studies will further define the role of MB as a targeted adjunct vasopressor.

CONCLUSION

Ongoing multicentre investigations, including pharmacokinetic-pharmacodynamic modelling studies (ClinicalTrials.gov NCT05941827), aim to individualize MB dosing using real-time hemodynamic monitoring. The Adjunctive MB in Septic SHock trial (NCT06900140) will assess feasibility prior to a large multicentre randomized trial, while additional studies are underway in Asia (NCT06481410, NCT0653224).

Accumulating evidence supports MB as a selective, mechanistically rational adjunct in septic shock. The most effective regimens employ early initiation (within 24 hours) and cumulative dosing up to approximately 4 mg/kg. Contemporary data demonstrate improvements in hemodynamic stability, vasopressor independence, myocardial performance, and renal perfusion without excess toxicity. Beyond septic shock, MB has utility in other forms of vasoplegia and selected toxicologic emergencies. Its low cost compared with newer vasopressors represents an additional advantage, particularly in low- and middle-income settings.