Role of nebulized heparin inhalation on mechanically ventilated critically ill patients

Introduction: Mechanical ventilation is one of the most important tools in the treatment of respiratory failure in critically ill patients, but it may cause lung injury and inflammatory response in the whole body. Aim of work: The aim of our study was to justify the effect of nebulized heparin on morbidity, oxygenation parameters, lung mechanics, and mortality in mechanically ventilated critically ill patients who are assumed to require mechanical ventilation for more than 48 h for different indications. Patients and methods: This study was conducted on 50 ICU patients who were in need of mechanical ventilation for more than 48 h. They were grouped randomly into two groups. One of the two groups was given nebulized heparin sodium until weaning or for a maximum of 14 days. Patients with coagulopathy or scheduled for any invasive intervention that may lead to bleeding were excluded. In addition, patients who were weaned or who died before day 4 of admission were also excluded. Both groups were followed up for a maximum of 28 days. The study medication was reduced or withheld if any significant bleeding occurred. The endpoint results were primary oxygenation parameters [mainly arterial oxygen partial pressure (PaO2)/inspired oxygen fraction (FIO2)] and ventilator-free days. All other data were recorded and analyzed to find out the adverse positive effect of heparin nebulization. Results: Data analysis revealed that the following data showed no statistically significant difference within groups over time or between the two groups: PaCO2, pH, PaO2, HCO3, SO2, PaO2/FIO2, FIO2×mean airway pressure/PaO2, peak inspiratory pressure, mean airway pressure, tumor necrosis factor α, systolic blood pressure, diastolic blood pressure, hemoglobin, hematocrit value, white blood cells count, platelet count, prothrombin concentration, international normalized ratio, presence of bloody sputum, ICU-free days at day 28, ventilator-free days at day 28, acute renal failure-free days at day 28, vasopressor-free days at day 28, and mortality and sputum culture results at day 4. However, the following data showed a statistically significant difference in the heparin-treated group: plateau pressure showed a statistically significant decrease between days 1 and 4 in the heparin-treated group (P=0.003) and a statistically significant difference when we compared the percentage change between the two groups (P=0.015). Compliance rate showed a statistically significant increase between days 1 and 4 in the heparin-treated group (P=0.019) but when we compared the percentage change between the two groups the difference was not statistically significant (P=0.256). Activated partial thromboplastin time showed a statistically significant increase between days 1 and 4 in the heparin-treated group (P=0.001), but when we compared the percentage change between the two groups the difference was not statistically significant (P=0.153). No cases of heparin-induced thrombocytopenia was noted in the heparin-treated group, nor was there major bleeding or need for blood transfusion related to the tested medication in this group. Conclusion: We recommend nebulized heparin as a safe drug that has a favorable effect in patients receiving mechanical ventilation especially with reduced compliance as in cases of acute respiratory distress syndrome.

the heparin-treated group: plateau pressure showed a statistically significant decrease between days 1 and 4 in the heparin-treated group (P=0.003) and a statistically significant difference when we compared the percentage change between the two groups (P=0.015). Compliance rate showed a statistically significant increase between days 1 and 4 in the heparin-treated group (P=0.019) but when we compared the percentage change between the two groups the difference was not statistically significant (P=0.256). Activated partial thromboplastin time showed a statistically significant increase between days 1 and 4 in the heparin-treated group (P=0.001), but when we compared the percentage change between the two groups the difference was not statistically significant (P=0.153). No cases of heparin-induced thrombocytopenia was noted in the heparin-treated group, nor was there major bleeding or need for blood transfusion related to the tested medication in this group.

Introduction
Mechanical ventilation is an important tool in the management of respiratory failure in the critically ill patient. It is required for the management of respiratory failure resulting from various clinical conditions such as acute respiratory distress syndrome (ARDS), pneumonia, sepsis, chronic obstructive pulmonary disease, and asthma. Although mechanical ventilation can be a lifesaving intervention, it is known to carry several side effects and risks [1]. Mechanical ventilation can increase the level of inflammatory mediators within the lungs, and treatment with the antagonists of these mediators may reduce it [2]. A number of potential targets have been identified in preclinical studies. Increased levels of several inflammatory mediators [including tumor necrosis factor (TNF) α , interleukin-6, and interleukin-10] were found in ex-vivo and in-vivo rat models subjected to injurious mechanical ventilation [3].
TNF has been consistently implicated in the pathogenesis of acute lung injury (ALI)/ventilatorinduced lung injury, both clinically and in experimental models [4]. In addition, lower TNF-α levels in both the serum and bronchoalveolar lavage fluid from patients at risk for ARDS exhibited a good negative predictive value for ARDS development [5].
Heparin and related molecules can bind electrostatically to the positively charged nuclear localization sequence of NF-κβ and prevent it from translocation to the nucleus. Blocking of this transcriptional factor can potentially reduce inflammatory gene activation and regulate the gene expression and production of proinflammatory cytokines, chemokines, and adhesion molecules [6].
Unfractionated heparin inhibits lipopolysaccharideinduced activation of endothelial cells through inhibition of p38MAPK and NF-κβ [7]. Heparin has been shown to bind to the surface of neutrophils and can inhibit their degranulation [8]. Further, heparin is able to inhibit neutrophil activation in response to thrombin-stimulated platelet products, in addition to inhibiting thrombin-induced platelets [9].

Aim of the work
The aim of our study was to justify the effect of nebulized heparin on the morbidity, oxygenation parameters, lung mechanics, and mortality of mechanically ventilated critically ill patients assumed to require mechanical ventilation for more than 48 h for different indications.

Patients and methods
This study was conducted in the respiratory ICU of New Kasr-Al-Aini Teaching Hospital and in the ICU of Beni-Suif University Hospital from October 2012 to January 2014. The study protocol was approved by local ethical committee and informed consent was taken .
Patients who had undergone mechanical ventilation for any indication and were assumed to require mechanical ventilation for more than 48 h were included in this study.

Exclusion criteria
Patients with the following criteria were excluded from our study.
(1) Patients who received mechanical ventilation for more than 24 h before enrollment.

Study design
This study was conducted on 50 ICU patients who were in need of mechanical ventilation for more than 48 h; they were selected and grouped randomly as follows.
(1) Group І: These patients were given nebulized heparin sodium until weaning or for a maximum of 14 days. (2) Group ІІ: These patients were considered as the comparative group.
Both groups were followed up for 28 days. Laboratory tests were followed up for the first 14 days only. The endpoint results were primary oxygenation parameters [mainly arterial oxygen partial pressure (PaO 2 )/ inspired oxygen fraction (FIO 2 )] and ventilator-free days. All other data were recorded and analyzed (on days 1 and 4) to find out the adverse/positive effect of heparin nebulization.
Heparin sodium at a dose of 25 000 U/5 ml was given by nebulization to group I in the inspiratory limb of the ventilator before the Y-piece every 4 h (150 000 U/ day). No dose adjustment was made for heparin administration for deep venous thrombosis prophylaxis, and nebulized heparin was given for a maximum of 14 days from randomization.

Weaning criteria
Spontaneous breathing trial was followed as the primary method of weaning, with other methods (e.g. gradual reduction of synchronized intermittent mandatory ventilation or pressure support titration) adopted as per the clinical condition. The decision to wean was taken on the basis of the following criteria.
(1) Resolution or stabilization of the underlying disease process. Criteria for a failed spontaneous breathing trial [10] Inadequate gas exchange.
(2) Hemodynamic instability. If any of these criteria were met, spontaneous breathing trial was terminated and the patient was placed back on previous ventilator settings for at least 24 h.
Patients were considered suitable for extubation if they were hemodynamically stable with an oxygen saturation of at least 95% while ventilated on pressure support less than or equal to 10 cmH 2 O, PEEP less than or equal to 5 cmH 2 O, and FIO 2 less than or equal to 50% and after passing the weaning parameters with intact cough reflex and need for endotracheal suction arising only at intervals greater than 4 h.
The following data were collected and subjected to statistical analysis.

Results
Data analysis revealed that the following data were not statistically significantly different within groups over time or when the percentage change between the two groups was compared: PaCO 2 , pH, PaO 2 , HCO 3 , SO 2 , PaO 2 /FIO 2 , FIO 2 ×mean airway pressure/PaO 2, peak inspiratory pressure, mean airway pressure, TNF α , systolic blood pressure, diastolic blood pressure, hemoglobin, hematocrit value, white blood cells count, platelet count, prothrombin concentration, INR, presence of bloody sputum, ICU-free days on day 28, ventilator-free days on day 28, acute renal failure-free days on day 28, vasopressor-free days on day 28, and mortality and sputum culture results on day 4.
However, the following data showed a statistically significant change in the heparin-treated group.  No statistically significant difference was found in TLC and PLT count between the two groups, nor on follow-up within each group. No statistical significant difference was found in hemoglobin and hematocrit between the two groups, nor on follow-up within each group. There was a statistical significant difference in aPTT on day 1 between the two groups, and on follow-up there was a statistically significant increase in aPTT in group I and decrease in group II. aPTT, activated partial thromboplastin time.  No statistically significant difference was found in PaO 2 and SpO 2 between the two groups, nor on follow-up within each group. No statistically significant difference was found in PaCO 2 and FIO 2 between the two groups, nor on follow-up within each group.  No statistically significant difference was found in PaO 2 /FIO 2 ratio between the two groups, nor on follow-up within each group. There was a statistically significant difference in all parameters on day 1 between the two groups, and on follow-up there was no statistically significant change. There was a statistically significant difference in plateau pressure on day 1 between the two groups, and on follow-up there was a statistically significant decrease in plateau pressure and increase in compliance rate in group I.
(1) Plateau pressure showed a statistically significant decrease between days 1 and 4 in the heparintreated group (P=0.003) and the difference was significant when we compared the percentage change between groups (P=0.015). (2) Compliance rate showed a statistically significant increase only between days 1 and 4 in the heparintreated group (P=0.019), but when we compared the percentage change between groups the difference was not statistically significant (P=0.256). (3) aPTT showed a statistically significant increase between days 1 and 4 in the heparin-treated group (P=0.001) but when we compared the percentage change in each group with each other this change was not statistically significant (P=0.153).
No cases of heparin-induced thrombocytopenia were noted in the heparin-treated group, nor was there major bleeding or need for blood transfusion related to the tested medication in this group.

Discussion
The current study was designed to assess the effects of nebulized heparin on the mechanically ventilated patient with respect to its safety and effects on morbidity and mortality (Tables 1,8 and 14).
A similar study conducted by Dixon et al. [11] was taken as a guide, with some modifications in the selection criteria (Dixon's study was performed on ALI patients, but our study was conducted on any patient expected to be mechanically ventilated for more than 48 h) and methodology.
The mean age of the studied patients was 56.24±14.1 years in group I versus 56.68±12.8 years in group II with a predominance of male sex in both groups (76 and 64% in groups I and II, respectively) without a statistically significant difference between the two groups (P=0.355). chronic obstructive pulmonary disease-associated respiratory failure was the predominant reason for mechanical ventilation in group I (36%), whereas pneumonia was the predominant reason in group II (28%). Radiographic   Hence, direct comparisons between the two groups at day 4 were avoided whenever possible.
Each group's data were compared between days 1 and 4 and the following was detected.
There was no statistically significant difference in either group in terms of arterial blood gas parameters (pH, PaCO 2 , PaO 2 , HCO 3 , and SpO 2 ) ( Tables 6 and 7). This matched the results of Li et al. [12], who found that there was no statistical difference in pH, PaO 2 , and PaCO 2 at the beginning versus the end of 5 h of mechanical ventilation in sheep given nebulized heparin. Our result also matched that of Dixon et al. [11], who found that there was no significant difference between groups in the partial pressure of arterial carbon dioxide over the course of the study period (days when the patients remained on mechanical ventilation to a maximum of 14 days from randomization).
Regarding other oxygenation parameters, in this study it showed no significant difference in either group (P The percentage change in the two groups was statistically significantly different in the following parameters: mean blood pressure, serum potassium, and plateau pressure. aPTT, activated partial thromboplastin time; Hb, hemoglobin; PEEP, positive end-expiratory pressure; PLT, platelet. value for PaO 2 /FIO 2 and FIO 2 ×mean airway pressure/ PaO 2 ) ( Table 9). This was in accordance with the results of Dixon et al. [13], who found that nebulized heparin did not cause significant changes in the ratio of PaO 2 to FIO 2 . In addition, Dixon et al. [11] found that the average daily PaO 2 /FiO 2 ratio while ventilated was similar in the nebulized heparin and nebulized placebo groups (194.2±62.8 vs. 187 ±38.6 mmHg, P=0.6). However, they found that, although not statistically significant, the PaO 2 /FiO 2 ratios were higher from day 3 in the heparin group, which did not match our findings (PaO 2 /FIO 2 was 230 ±140.8 on day 1 vs. 206.6±68.5 on day 4 in group I and 204.6±92.4 vs. 240.4±87.2 in group II) ( Table 9).
In contrast, Murakami et al. [14] found that the PaO 2 / FiO 2 ratio dropped markedly in saline-treated sheep with induced ARDS. However, in the heparinnebulized group, the drop was significantly attenuated after 12 h. The pulmonary shunt fraction increased in the saline-treated group, reaching 50-60% at 24 h, but was significantly lower in the heparinnebulized group.
In the current study, there were no statistically significant changes in either group regarding peak inspiratory pressure and mean airway pressure (Table 10).
When we assessed plateau pressure and compliance, we found a statistically significant decrease in plateau pressure and increase in compliance in group I (the heparin-treated group). This statistically significant change was not found in group II (Table 11). In contrast, Dixon et al. [13] found that there was no statistically significant change in lung compliance for the dosage, nor in the interaction between dosage and time.
TNF was tested before and after treatment with nebulized heparin in group I but no statistically significant difference was noted (Table 12). The same result was seen when we compared the two groups on day 4 (Table 13). This was similar to the findings of Hofstra et al. [15], who stated that bronchoalveolar levels of TNF and histopathology of the lungs were not affected by nebulization with anticoagulants.
This finding also matched that of Dixon et al. [11], who noted that levels of TNF in pulmonary lavage fluid were similar in the two groups at baseline and on each day that samples were taken following enrollment in the groups (with nebulized heparin and placebo).
Regarding systolic blood pressure, we noted an increase in mean value from days 1 to 4 in group I only (106±22 mmHg on day 1 and 112±31 mmHg on day 4 in group I vs. 118±25 mmHg on day 1 and 118±24 mmHg on day 4 in group II), but these changes were not statistically significant (P=0.156 and 0.833, respectively).
Diastolic blood pressure showed an increase in mean value from days 1 to 4 in both groups (68±14 mmHg on day 1 and 70±18 mmHg on day 4 in group I vs. 74±17 mmHg on day 1 and 75±14 mmHg on day 4 in group II), but this increase was not statistically significant (P=0.127 and 0.932, respectively).
However, when the mean arterial blood pressure was analyzed we noticed an increase in mean blood pressure in group I only (79±15 mmHg on day 1 and 84±22 mmHg on day 4), which was statistically significant (P=0.022), whereas group II did not show that increase (89±19 mmHg on day 1 and 89±16 mmHg on day 4) (P=0.888) ( Table 15).
This contradicted the results of Murakami et al. [16], who noted that the mean arterial pressure decreased and was not attenuated by intravenous heparin administration in sheep with induced ALI.
Comparison of hemoglobin, hematocrit value, white blood cells, platelet count, and INR showed no statistically significant difference between days 1 and 4 in the two groups (Tables 2-4). There was no need for blood transfusion related to the drug tested.
Yip et al. [17] tested the effect of nebulized heparin at a dose of 30 000 U/day on burn patients with inhalation lung injury; they noted that platelet count and prothrombin time followed a similar trend in both groups (nebulized heparin with N-acetyl cystine or salbutamol group and control group). In addition, there were no statistical differences in the mean platelet counts of the treatment and the control group over the 7 days.
Regarding aPTT there were no statistically significant differences in group II. However, in group I, we noted a statistically significant increase in aPTT (Table 5) but this increase did not reach the therapeutic level. This matched the results of Dixon et al. [13], who monitored the anticoagulation effect of nebulized heparin at doses of 50 000, 100 000, 200 000, and 400 000 U/day and found no serious adverse events for any dose.
Bloody sputum incidence was 28% in group I versus 36% in group II. Analysis revealed that there was no statistical significance for this difference (P=0.544). No frank bleeding or marked bleeding tendency was noted in either group. This was similar to the findings of Dixon et al. [13], who have not recorded any serious adverse event in their patients except for one patient in the 400 000 U/day group who developed blood-stained respiratory secretions after the seventh dose. In addition, Ahmed et al. [18] failed to demonstrate any anticoagulant effect of inhaled heparin on bleeding tendency in his trial on patients with bronchial asthma. This was confirmed by Dixon et al. [11].
No cases of heparin-induced thrombocytopenia was reported in any of our patients, matching the results of Yip et al. [17].
The mean number of ICU-free days was 6±9 days in group I versus 4±8 days in group II. Group I showed more number of ICU-free days but this was not statistically significant (P=0.433) even after omitting nonsurvivors (P=0.354) ( Table 13). Our results were similar to those of Dixon et al. [11], who found that the duration of ICU stay was 9.4±7.4 in the heparintreated group versus 14.0±13.1 days in the control group (P=0.2). In addition, Yip et al. [16] showed that the median length of ICU stay was 6 days in the heparin-treated group compared with 7 days in the control group.
The mean number of ventilator-free days was 10±11 days in group I versus 7±10 days in group II. Group I showed more ventilator-free days but this was not statistically significant (P=0.266) even after omitting nonsurvivors (P=0.287) (Table 13). When we compared the percentage of weaning in each group (40% in group I vs. 28% in group II), it was higher in group I but this was not statistically significant (P=0.37) even after omitting nonsurvivors (P=0.492). This result was similar to that of Yip et al. [17], who showed that the median duration of intubation was 5 days in both groups.
In contrast, Dixon et al. [11] found that nebulized heparin was associated with improved number of ventilator-free days among survivors at day 28 (22.6 ±4.0) in the heparin-treated group versus 18.0±7.1 in the control group (P=0.02).
Mean acute renal failure-free days was 23±10 days in both groups, and this was not statistically significant (P=0.978) even after omitting nonsurvivors (P=0.317) ( Table 13). This result was similar to that of Dixon et al. [11], who found that the average number of renal failure-free days among survivors at day 28 (28 days in both groups) showed no statistically significant difference between the two groups (P=0.09).
Mean vasopressor-free days was 21±11 days in group I versus 22±11 days in group II, and this was not statistically significant (P=0.773) even after omitting nonsurvivors (P=0.951) ( Table 13). Our results were similar to those of Dixon et al. [11], who found that the number of vasopressor-free days among survivors at day 28 was 24.7(±3.2) in the heparin-treated group versus 22.0±7.0 days in the control group (P=0.12).
The number of deaths in both groups was eight persons (32%) (no statistically significant difference; P=1) ( Table 16). This was similar to the finding of Dixon et al. [11], who found that mortality at day 28 was similar in the two groups (P=0.7).

Limitations in the current study
(1) The sample size for the groups is small.
(2) The incidence of bleeding and other complications reported in the study depended mostly on the documentation of nurses and other physicians.

Conclusion
From the current study, we conclude the following.
(1) Heparin at a dose of 100 000 to 150 000 U/day is a safe drug when used through nebulization and does not affect coagulation markedly. (2) Nebulized heparin improved the compliance rate and significantly reduced the plateau pressure in our tested patients. (3) Nebulized heparin did not affect oxygenation, mortality, vasopressor-free days, incidence of acute renal failure, prothrombin concentration, hemoglobin concentration, and platelet count in the tested group. (4) The number of ventilator-free days was higher in the heparin-treated group but did not reach a statistically significant value.

Recommendations
Nebulized heparin is a safe drug that has a favorable effect in patients receiving mechanical ventilation, especially with reduced compliance as in ARDS.
Repetition of this study on a larger scale would verify its results as there are no available studies on a larger number of patients.
(1) Nebulized heparin should be tested on specific indications of mechanical ventilation to detect the effect of heparin on individual disease.
A similar study should be conducted over a longer follow-up period as it may show a statistically significant difference regarding ventilator-free days and mortality.