INTRODUCTION:

The elderly population is a group that is vulnerable to severe complications due to influenza infection¹. This condition is caused by immunosenescence, a decreased immune system function with age. Immunosenescence leads to a delayed and less efficient immune response against pathogens like influenza viruses². Vaccination is becoming essential in protecting this population from influenza infection, which can lead to high morbidity and mortality³. Influenza vaccination in the elderly population is important for several scientific reasons⁴. Immunosenescence, the decline in immune function with age, makes the elderly more susceptible to infection and reduces their ability to fight pathogens such as influenza viruses⁵. The elderly are also at high risk of severe complications due to influenza, such as pneumonia, bronchitis, exacerbations of chronic diseases, and death. Data shows most influenza-related deaths occur in individuals aged 65 years and older, and serious complications often require intensive care that increases health costs⁶.

Although the elderly immune response to vaccines is lower than in the younger population, influenza vaccination still provides significant protection by reducing influenza incidence, hospitalization, and mortality. Vaccination also contributes to herd immunity, protecting individuals who cannot be vaccinated or have a weak immune response⁴. This helps reduce the spread of the virus within the population and protects the wider community. Influenza vaccination is an effective and affordable prevention strategy that reduces the incidence and severity of illness and related complications in the aging population⁷. The trivalent influenza vaccine is designed to protect against the three main strains of the influenza virus, hoping to boost immunity and reduce the risk of infection and related complications⁸. Thus, influenza vaccination not only protects the aging populationfrom the risk of infection and serious complications but also reduces the public health burden, improves the health and quality of life of older people, and maintains the stability of the health system⁹.

The immunological markers IL-15, MIP-1α, IFN-γ, and CD16 are essential in the immune response to influenza infection¹⁰. IL-15 and CD16 play a role in NK cell activation and function, while MIP-1α is involved in the recruitment of immune cells to the site of infection, and IFN-γ plays a role in mediating cellular responses¹¹. Understanding how these markers interact during influenza infection could help develop more effective vaccination strategies and therapeutic targets for enhancing immune responses in elderly populations¹². A better understanding of how these vaccines modulate immune responses could aid in developing more effective vaccination strategies to protect elderly populations from influenza¹³.

This study is crucial for the health of older adults as it examines how the trivalent influenza vaccine affects their immune responses. The study provides insights into how vaccines modulate humoral and cellular immunity by evaluating immunological parameters like IL-15, MIP-1α, IFN-γ, and CD16. The findings highlight the variability of immune responses in seniors, potentially informing more effective and personalized vaccination strategies. The study shows the vaccine's effectiveness in boosting antibody titers and altering immunological markers, underscoring the importance of influenza vaccination in reducing morbidity and mortality among the elderly.

MATERIAL AND METHODS:

This research applies the ethical principles of storing biological materials and has been approved by the research ethics committee of Padjadjaran University with approval number 88/UN6. KEP/EC/2019. The design of this study is a cohort study with analytical observational research methods that examine the relationship between immunological marker responses after vaccination with trivalent influenza vaccine in 44 elderly populations using intervariable analysis methods.

Subject Selection:

The subjects of the study were selected based on several criteria. Subjects must be 60 years or older, in good health, or have stable chronic disease, and give written consent after receiving a full study description. Subjects with severe allergies to vaccine components, acute illness, immunodeficiency conditions, have received influenza vaccine in the past six months, or have other severe medical conditions will be excluded. Initial screening is done through an interview and medical history examination.Subjects who meet the criteria then undergo a complete medical assessment. After giving written consent, subjects were randomly divided into different vaccine groups or received a combined trivalent vaccine. During the study, subjects were monitored to ensure their safety and collect the necessary data. The entire selection process follows the ethical guidelines of medical research and has received approval from the relevant ethics committee.

Serum Collection:

Blood sampling begins with the vein area cleaned using an alcohol swab, a tourniquet is placed on the upper arm, and a syringe is carefully inserted to collect 5-10 mL of blood into the vacutainer tube. The tourniquet is then removed, the syringe is removed, the injection area is pressed with alcohol cotton, and plaster is applied. The tubes are labeled with subject information, date, and time of retrieval. Sampling after vaccination for one month is carried out using the same procedure. Blood sample processing is performed with a centrifuged blood tube at 3000rpm for 10-15 minutes to separate the serum. The formed serum is transferred to a clean micro-tube using a sterile pipette, and then the serum is frozen and stored at -80°C until analysis¹⁴.

ELISA Assay:

The ELISA method for examination of immunological markers IL-15, MIP-1α, IFN-γ, and CD16 in blood serum is carried out through several steps. Blood serum is collected and stored at -80°C. ELISA plates are coated with specific arresting antibodies, incubated overnight at four °C, and then washed with a wash buffer. The diluted or standard serum is added to the plate wells and incubated for two hours at room temperature, followed by washing. Detection antibodies are added and incubated for 1-2 h, followed by re-washing. An enzyme substrate (TMB) is added and incubated for 20-30 min in a dark place, and the reaction is stopped with 2N sulfuric acid. The absorbance is read at 450 nm, and a standard curve determines the marker concentration. Positive and negative controls were used to validate the results, and all reagents were at room temperature before use. This method allows accurate measurement of serum immunological marker concentrations to evaluate the immune response to trivalent influenza vaccine in the elderly population¹⁵.

Statistical Analyses:

Statistical analysis in this study was conducted using the Kruskal-Wallis test, Wilcoxon Signed-Rank, and Spearman correlation with a meaning limit of p<0.05. The Kruskal-Wallis assay showed significant differences in the titers of HAI and IFN-γ antibodies among groups receiving different trivalent vaccines but not in IL-15 and MIP-1α. The Wilcoxon Signed-Rank test showed substantial changes in HAI and IFN-γ antibody titers before and after vaccination. The Spearman correlation suggests that the association between HAI antibody titers and immunological markers is weak and insignificant.

RESULTS:

This study evaluated the immunological response to trivalent influenza vaccine in the elderly population, focusing on several immunological biomarkers and their correlation with antibody reaction. In addition, the study explored the relationship between sex, age factors, and comorbid diseases in the elderly population.

Table 1 shows the distribution of comorbid subjects by sex and age, with comorbidities <4 and ≥4. Of the 44 subjects, 95.5% had <4 comorbidities, and only 4.5% had ≥4 comorbidities. By gender, males comprised 15.9% of subjects (13.6% with comorbidities <4 and 2.3% with comorbidities ≥4), while females comprised 84.1% of subjects (81.8% with comorbidities <4 and 2.3% with comorbidities ≥4). Statistical analysis showed no significant association between sex and comorbidity (p= 0.177). By age, subjects aged 60-69 years comprised 59.1% (56.8% with comorbidities <4 and 2.3% with comorbidities ≥4), subjects aged 70-79 years comprised 38.6% (36.4% with comorbidities <4 and 2.3% with comorbidities ≥4), and subjects aged <80 years comprised 2.3% with comorbidities <4. There was no significant association between age and comorbidity (p = 0.910).

Table 2. Distribution of subjects by type of comorbidity based on gender and age

S. No	Comorbid	Amount	%	Sex				Ages (Years)
				Male		Female		60-69		70-79		< 80
				n	%	n	%	n	%	n	%	n	%
1	Hypertension	25	38,46	3	4,62	22	33,85	12	18,46	13	20,00	0	0,00
2	Gout	8	12,31	1	1,54	7	10,77	6	9,23	2	3,08	0	0,00
3	Cholesterol	5	7,69	1	1,54	4	6,15	4	6,15	1	1,54	0	0,00
4	Gastritis	5	7,69	0	0,00	5	7,69	3	4,62	2	3,08	0	0,00
5	Osteoarthritis	5	7,69	1	1,54	4	6,15	4	6,15	1	1,54	0	0,00
6	Vertigo	3	4,62	1	1,54	2	3,08	1	1,54	2	3,08	0	0,00
7	Strokes	2	3,08	1	1,54	1	1,54	1	1,54	1	1,54	0	0,00
8	Rheumatoid	2	3,08	1	1,54	1	1,54	0	0,00	2	3,08	0	0,00
9	Arthritis	2	3,08	0	0,00	2	3,08	1	1,54	1	1,54	0	0,00
10	Dyslipidemia	2	3,08	0	0,00	2	3,08	2	3,08	0	0,00	0	0,00
11	GERD	2	3,08	1	1,54	1	1,54	1	1,54	1	1,54	0	0,00
12	TB_Lung	1	1,54	1	1,54	0	0,00	1	1,54	0	0,00	0	0,00
13	BPH	1	1,54	1	1,54	0	0,00	0	0,00	1	1,54	0	0,00
14	Toothache	1	1,54	0	0,00	1	1,54	0	0,00	1	1,54	0	0,00
15	Osteoporosis	1	1,54	0	0,00	1	1,54	0	0,00	1	1,54	0	0,00

GERD (Gastroesophageal Reflux Disease), TB (pulmonary tuberculosis) and BPH (Benign Prostatic Hyperplasia)

Table 2 shows the distribution of subjects by comorbid disease type, sex, and age. Hypertension was the most common comorbidity, experienced by 38.46% of subjects, with 4.62% of males and 33.85% females, more in the age groups of 60-69 years (18.46%) and 70-79 years (20.00%). Uric acid (12.31%) is more common in women (10.77%) and the age group of 60-69 years (9.23%). High cholesterol (7.69%) and osteoarthritis (7.69%) were more common in the age group of 60-69 years. Other diseases such as vertigo, stroke, rheumatism, arthritis, dyslipidemia, and GERD were experienced by 3.08% to 4.62% of subjects, respectively. Diseases such as pulmonary TB, BPH, toothache, and osteoporosis were each found in 1.54% of subjects, with pulmonary TB and BPH more common in males and osteoporosis only in women. Most comorbid diseases are found in 60-69 and 70-79 years, without comorbidities, in subjects aged <80.

Table 3 reports that Hemagglutination-Inhibition (HI) values are used to measure antibody levels against influenza viruses, where higher values indicate better protection. The results showed that before vaccination, the average HI value for A/Singapore – EFTHA 1835 was 204.55 ± 27.15, for A/Singapore – EFTHAS 1828 was 145.00 ± 27.7, and for B/Maryland – EFTTHAC 1822 was 41.82 ± 4.51. After vaccination, the mean HI scores increased significantly to 1044.09±191.22 for A/Singapore – EFTHA 1835, 463.64±71.02 for A/Singapore – EFTHAS 1828, and 289.55±49.53 for B/Maryland – EFTTHAC 1822, with all increases having p-value = 0.000, indicating a very significant change.

The A/Singapore – EFTHA 1835 vaccine showed the highest increase in HI values, about five times the initial value, providing strong protection against influenza viruses and making it the most influential vaccine among the three tested vaccines. The A/Singapore vaccine – EFTHAS 1828 also showed a significant increase, more than tripling, providing reasonably good protection. Meanwhile, the B/Maryland vaccine – EFTTHAC 1822, although started with lower HI values, increased almost sevenfold after vaccination, providing adequate protection. Overall, all three influenza vaccines effectively increased antibody titers and protected against influenza viruses in the elderly, with A/Singapore – EFTHA 1835 being the superior.

Table 4 shows the results of Spearman's rho correlation analysis between A/Singapore – EFTHA 1835 vaccine response and various immunological markers (IL-15, MIP-1α, IFN-γ, and CD16) in the elderly population. For this vaccine, no significant correlation was found with immunological markers tested. The correlation coefficient for IL-15 is 0.197(p-value = 0.199), for MIP-1α it is 0.178 (p= 0.249), for IFN-γ it is -0.02 (p= 0.899), and for CD16 it is 0.221(p= 0.149). These results suggest that changes in HAI values for the A/Singapore – EFTHA 1835 vaccine were not associated with these markers, so the immune response to these vaccines did not significantly affect the immunological markers tested.

Correlation analysis for the A/Singapore – EFTHAS 1828 vaccine showed that only CD16 significantly correlated with vaccine response. The correlation coefficient for CD16 was 0.373 with a p-value of 0.013, indicating a significant positive association between vaccine response and increased CD16 expression. The A/Singapore vaccine – EFTHAS 1828 can better stimulate NK cell activation. However, other markers such as IL-15, MIP-1α, and IFN-γ showed no significant correlation with correlation coefficients of -0.054 (p = 0.73),-0.014(p=0.927), and -0.265(p=0.082), respectively. Therefore, this vaccine response was not associated with significant changes in these markers other than CD16.

The B/Maryland vaccine – EFTTHAC 1822 also showed a significant positive correlation with CD16, with a correlation coefficient of 0.323 and a p-value of 0.033. This suggests that this vaccine response is associated with increased CD16 expression, which indicates NK cell activation. However, no significant correlations were found for markers IL-15, MIP-1α, and IFN-γ, with correlation coefficients of 0.112 (p = 0.469), -0.189 (p=0.218), and 0.109 (p = 0.483), respectively. These results suggest that although the B/Maryland vaccine – EFTTHAC 1822 affected NK cell activation, the immune response to this vaccine was not associated with significant changes in IL-15, MIP-1α, and IFN-γ markers.

Table 5 shows statistical analysis of the response of Hemagglutination Inhibition (HAI) values of three types of influenza vaccines (A/Singapore – EFTHA 1835, A/Singapore – EFTHAS 1828, and B/Maryland – EFTTHAC 1822) to various immunological markers (IL-15, MIP-1α, IFN-γ, and CD16) in the elderly population before and after vaccination. Before vaccination, the highest HAI titer was recorded in the A/Singapore – EFTHA 1835 vaccine (204.55±27.15), followed by A/Singapore – EFTHAS 1828 (145.00± 27.7), and the lowest in B/Maryland – EFTTHAC 1822 (41.82±4.51). Average concentrations of IL-15, MIP-1α, IFN-γ, and CD16 were the same for all three vaccines, at 5.786±0.915pg/mL, 113.356±26.052pg/mL, 2.024± 0.228pg/mL, and 31,034±7,094pg/mL, respectively. After vaccination, there was a significant increase in HAI titers for all vaccines, with the most significant increase being A/Singapore – EFTHA 1835 (1044.09± 191.22), followed by A/Singapore – EFTHAS 1828 (463.64±71.02), and B/Maryland – EFTTHAC 1822 (289.55±49.53), with a p=0.000. This indicates significant differences between vaccines before and after vaccination.

For IL-15, after vaccination, concentrations increased to 6.581±2.472pg/mL for all three vaccines, although no p-value was given to compare these specific changes. MIP-1α concentrations also increased to 132.992± 29.298pg/mL after vaccination for all three vaccines, but again, no p-value was given to compare these changes. Before vaccination, the mean concentration of IFN-γ was 2.024±0.228pg/mL for all three vaccines. After immunization, this concentration decreased to 1.027± 0.108pg/mL, suggesting that vaccination significantly lowered IFN-γ levels. Finally, for CD16, before immunization, mean concentrations were 31.034±7.094 pg/mL for all three vaccines. After immunization, concentrations decreased to 22.777±2.840pg/mL, suggesting vaccination significantly lowers CD16 levels. This table shows that trivalent influenza vaccines significantly increase HAI titers and affect levels of several immunological markers, particularly IFN-γ and CD16, in the elderly. These significant changes in HAI titers and immunological markers indicate the vaccine's effectiveness in stimulating an immune response in this population.

DISCUSSION:

This study evaluated the effectiveness of the trivalent influenza vaccine in enhancing immune responses in elderly populations by measuring immunological parameters such as IL-15, MIP-1α, IFN-γ, and CD16. The elderly population is susceptible to influenza infection and serious complications due to decreased immune system function. The study analyzed changes in antibody titers and immunological markers before and after vaccination. Results showed significant differences in HAI and IFN-γ antibody titers and substantial changes before and after vaccination. The correlation between HAI antibody titers and immunological markers showed a weak and insignificant association. The discussion will outline the interpretation of the results, clinical implications, limitations of the study, and contributions to more effective vaccination strategies for the elderly population, as well as the need for further research on booster doses and long-term immune responses.

Table 1 reports the distribution of comorbid subjects by sex and age and their relationship to trivalent influenza vaccination in the elderly population. The results of this study are consistent with previous studies showing that comorbidities are more common in the elderly population but are not necessarily related to a specific gender or age range¹⁶. In this study, although the proportion of women was higher than men, no significant association was found between sex and comorbid rates. This aligns with the findings by Canuto et al. (2018), who also found no significant differences in comorbidity rates by sex in the elderly population¹⁷. In addition, the study also showed that age was not significantly correlated with comorbid rates, which supports previous findings by Thavorn et al. (2017), where variations in comorbidities were not directly related to age. Instead, it was influenced by other factors such as lifestyle and access to health care¹⁸. Although the elderly tend to have more chronic diseases, these results suggest that the increase in comorbidities does not necessarily go hand in hand with the increase in age.

Table 2 reports the distribution of subjects by comorbid type by sex and age. These results are consistent with previous studies showing that hypertension is one of the most common comorbidities in the elderly population¹⁹. A higher prevalence of hypertension in women has also been documented in other studies²⁰. Gout is more commonly found in elderly females, which aligns with findings that gout becomes more common after menopause²¹. High cholesterol and osteoarthritis are common comorbidities in the age group of 60-69 years, and these conditions are also supported by studies showing an increase in the prevalence of these conditions²².

The prevalence of other diseases such as vertigo, stroke, rheumatism, arthritis, dyslipidemia, and GERD (Gastroesophageal Reflux Disease) in study subjects is also in line with the existing literature, which suggests that these diseases are often found in the elderly population²³. TB (pulmonary tuberculosis) and BPH (Benign Prostatic Hyperplasia) are more common in males according to the epidemiological profile of the disease, where pulmonary TB is often found in the elderly male population, and BPH is a common condition in older men²⁴. Osteoporosis, which is only found in women, also supports previous research showing that osteoporosis is more common in postmenopausal women²⁵.

Table 3 reports evaluating the effectiveness of trivalent influenza vaccine in enhancing immune responses in elderly populations through measurement of Hemagglutination-Inhibition (HI) values, which are used to measure antibody levels against influenza viruses.Higher HI values indicate better protection against infection. Immunosenescence is a decline in immune system function that occurs with age. This causes the elderly population to be more susceptible to infections, including influenza, and a weaker response to vaccination¹³. The study showed that despite age-related declines in immune function, trivalent influenza vaccines were able to increase antibody titers in the elderly population significantly. The A/Singapore – EFTHA 1835 vaccine showed the highest increase in HI values, about five times the initial value, providing strong protection against influenza viruses and making it the most effective vaccine among the three tested vaccines. These results align with previous studies showing that more immunogenic vaccines can partially overcome the effects of immunosenescence and provide better protection in elderly populations²⁶.

The A/Singapore vaccine – EFTHAS 1828 also showed a significant increase, more than tripling, providing fairly good protection. Although not as strong as the A/Singapore – EFTHA 1835 vaccine, this increase shows that this vaccine is still effective in enhancing the immune response in the elderly. This supports the finding that different vaccine formulations can provide various levels of protection but still benefit the elderly population (Nikolich-Žugich, 2008). Meanwhile, the B/Maryland vaccine – EFTTHAC 1822, although started with lower HI values, increased almost sevenfold after vaccination, providing adequate protection. This suggests that although the vaccine is initially less immunogenic, it can stimulate a significant immune response after vaccination, following the literature stating that even a low initial reaction can be substantially improved through appropriate vaccination ²⁷.

Table 4 evaluates the correlation between trivalent influenza vaccine response and correlations of various immunological markers (IL-15, MIP-1α, IFN-γ, and CD16) in elderly populations. The decline in immune function with age includes changes in immune cells, such as T, B, and NK cells, all of which play an essential role in response to vaccination²⁸. The results of this study showed that although the reaction to the A/Singapore – EFTHA 1835 vaccine was not significantly correlated with the immunological markers tested, it could still substantially increase the HAI value. This supports the finding that humoral responses can be maintained despite age-related declines in cellular function²⁹. In contrast, the A/Singapore – EFTHAS 1828 and B/Maryland – EFTTHAC 1822 vaccines significantly correlated with CD16, indicating that both vaccines can stimulate NK cell activation, vital in the immune response to influenza viruses.

Elevated CD16 indicates NK cell activation, which may help compensate for decreased T and B cell function in the elderly population³⁰. NK cells control viral infections through cytotoxic mechanisms and cytokine production ³¹. Therefore, vaccines that increase NK cell activation may provide additional protection for the elderly. However, the absence of significant correlation with markers IL-15, MIP-1α, and IFN-γ suggests that the immune response to these vaccines does not depend on just one immunological pathway but rather results from a complex interaction of various immune system components. It emphasizes the importance of considering a multifaceted approach in vaccine development and evaluation for the elderly population.

Table 5 evaluates the effectiveness of three types of trivalent influenza vaccines (A/Singapore – EFTHA 1835, A/Singapore – EFTHAS 1828, and B/Maryland – EFTTHAC 1822) in enhancing immune responses in the elderly population, focusing on changes in Hemagglutination Inhibition (HAI) titers and various immunological markers (IL-15, MIP-1α, IFN-γ, and CD16). Age-related decline in immune function is a major challenge in vaccination in older people, reducing the effectiveness of their immune response to vaccines ³². Before vaccination, the highest HAI titer was recorded in vaccine A/Singapore – EFTHA 1835, followed by A/Singapore – EFTHAS 1828, and lowest in B/Maryland – EFTTHAC 1822. After vaccination, all three vaccines showed a significant increase in HAI titers, with the largest increase being A/Singapore – EFTHA 1835. This increase suggests that all three vaccines effectively stimulate antibody responses in the elderly population, which is particularly important given the decreased ability of humoral immune responses in the elderly due to Immunosenescence³³. The A/Singapore – EFTHA 1835 vaccine showed the highest increase, suggesting it may be more immunogenic and effective in this population.After vaccination, there was an increase in IL-15 and MIP-1α concentrations for all three vaccines. IL-15 and MIP-1α are important cytokines in the immune response, with IL-15 playing a role in the proliferation of T and NK cells and MIP-1α in immune cell chemotaxis³⁴. In contrast, IFN-γ concentrations decreased significantly after vaccination. IFN-γ is a pro-inflammatory cytokine important in response to viral infections³⁵. This decrease may indicate that vaccination helps reorganize cytokine balance, reducing the excessive inflammatory response often associated with Immunosenescence³⁶.

CD16 concentrations also decreased after vaccination. CD16 is a marker of NK cell activation, and this decrease may indicate that vaccination reduces NK cell overactivity, which may be a sign of a more balanced immune system reset³⁷. This decrease is consistent with research showing that modulation of NK cell activity is essential in effective immune responses in the elderly³⁰. Immunosenescence leads to alterations in immune cells, such as T, B, and NK cells, which play crucial roles in the response to vaccination³⁸. The study showed that the trivalent influenza vaccine significantly increased HAI titers despite immunosenescence, suggesting that humoral responses can still be effectively stimulated. The decrease in IFN-γ and CD16 suggests that vaccination may help reorganize the inflammatory response, essential for preventing excessive tissue damage in older adults³⁹.

Table 6 evaluates the immunological response to trivalent influenza vaccine in the elderly population. The mean concentration of IL-15 before vaccination was 5,786±0.915pg/mL, which increased to 6,581±2,472 pg/mL after immunization. Despite the increase, IL-15 is essential in NK and T cells' proliferation and activity⁴⁰. The insignificance of these changes suggests that trivalent influenza vaccines may not be potent enough to stimulate a significant increase in IL-15 production in the elderly population, which may be due to a decrease in the ability of immune cells to respond to stimulation due to immunosenescence.Meanwhile, MIP-1α concentrations were 113,356±26,052pg/mL, increasing to 132,992±29,298 pg/mL after vaccination. MIP-1α is a chemokine that recruits immune cells to the site of infection⁴¹. It can be assumed that these changes suggest that the chemokine response to vaccination in the elderly may not be sufficient to overcome immunosenescence-induced functional decline, indicating that vaccines may not increase immune cell mobilization in this population optimally. Results from IFN-γ markers show different results. Before vaccination, the average IFN-γ concentration was 2,024±0.228pg/mL, which decreased to 1,027±0.108pg/mL after vaccination. This decrease was statistically significant (p-value = 0.003). IFN-γ is a pro-inflammatory cytokine important in the immune response to viral infections ⁴². This significant decrease may indicate that vaccination helps re-regulate cytokine balance in the elderly, reducing the exaggerated inflammatory response often associated with Immunosenescence⁴³. This can be a positive sign that vaccination can help reduce chronic inflammation usually experienced by the elderly.In addition to CD16 titers, the mean concentration before vaccination was 31,034±7,094pg/mL, which decreased to 22,777 ±2,840 pg/mL after vaccination. CD16 is a marker of NK cell activation, and this decrease may indicate that vaccination reduces NK cell overactivity. NK cells control viral infections through cytotoxic mechanisms and cytokine production³¹. The insignificance of these changes could suggest that despite some modulation, vaccination does not dramatically alter NK cell activity in the elderly population.

Table 7 analyzed the correlation between four immunological markers (IL-15, MIP-1α, IFN-γ, and CD16) in the elderly population after administration of trivalent influenza vaccine using analysis. This study showed no significant correlation between immunological markers tested after trivalent influenza vaccination in the elderly. This indicates that a change in one immunological marker is not significantly associated with a change in another. These findings are relevant in immunosenescence, where immune responses in elderly populations are often fragmented and poorly coordinated⁴⁴. This study underscores the complexity of immune responses in older people, where decreased coordination between different components of the immune system can reduce the effectiveness of vaccination⁴⁵. Immunosenescenceminimizes the number of immune cells and interferes with the function and communication between immune cells, which may explain why no significant correlation was found between the markers tested⁴⁶. Previous studies have shown that older adults often experience a decline in adaptive immune responses, including decreased production of pro-inflammatory cytokines and less effective cellular activity⁴⁷. In this context, the results of this study indicate that although trivalent influenza vaccines may mount specific immune responses, as seen in increased antibody titers, overall coordination in the immune response may remain fragmented in the elderly population.

This study showed that trivalent influenza vaccines could increase antibody titers in the elderly. However, no significant correlation existed between changes in immunological markers IL-15, MIP-1α, IFN-γ, and CD16. These results underscore the complexity of the immune response in the elderly due to immunosenescence, where fragmented immune reactions can reduce the effectiveness of vaccination. The findings emphasize the need for more research to understand the mechanisms underlying the immune response in the elderly population and develop more effective vaccination strategies to improve immune response coordination and protection against influenza.

CONCLUSION:

This study showed that all three types of trivalent influenza vaccines (A/Singapore – EFTHA 1835, A/Singapore – EFTHAS 1828, and B/Maryland – EFTTHAC 1822) were effective in increasing the titer of Hemagglutination Inhibition (HAI) in the elderly population, with the most significant increase in the A/Singapore – EFTHA 1835 vaccine. Although there was an increase in IL-15 and MIP-1α concentrations, a considerable decrease in IFN-γ, and a reduction in CD16 after vaccination, these changes were not statistically significant except for IFN-γ. These results highlight the challenge of immunosenescence reducing the effectiveness of the immune response in the elderly but still show that trivalent influenza vaccination canmount the humoral immune response effectively.

CONFLICT OF INTEREST:

The authors declare no conflicts of interest.

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Received on 22.05.2024 Revised on 16.09.2024

Accepted on 06.12.2024 Published on 02.05.2025

Available online from May 07, 2025

Research J. Pharmacy and Technology. 2025;18(5):2281-2290.

DOI: 10.52711/0974-360X.2025.00327

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Creative Commons License.

Treatments	Vaccine Types	Titer HAI (HI Unit)	N	Trivalent Vaccine				*p-value*
				Mean±SD (pg/mL)
				IL-15	MIP 1 α	IFN γ	CD16
Pree Vaccination	A/Singapore – EFTHA 1835	204.55± 27.15	44	5.786± 0.915	113.356± 26.052	2.024± 0.228	31.034± 7.094	0.000**
	A/Singapore – EFTHAS 1828	145.00± 27.7	44	5.786 ± 0.915	113.356 ± 26.052	2.024 ± 0.228	31.034± 7.094
	B/Maryland –EFTTHAC 1822	41.82± 4.51	44	5.786 ± 0.915	113.356 ± 26.052	2.024 ± 0.228	31.034 ± 7.094
	p-value	0,000*
Post Vaccination	A/Singapore – EFTHA 1835	1044.09± 191.22	44	6.581 ± 2.472	132.992 ± 29.298	1.027 ± 0.108	22.777 ± 2.840
	A/Singapore – EFTHAS 1828	463.64± 71.02	44	6.581 ± 2.472	132.992 ± 29.298	1.027 ± 0.108	22.777 ± 2.840
	B/Maryland –EFTTHAC 1822	289.55± 49.53	44	6.581 ± 2.472	132.992 ± 29.298	1.027 ± 0.108	22.777 ± 2.840
	p-value	0,000*

Treatments	N	Mean±SD (pg/mL)				*p-Value
Treatments	N	IL-15	MIP 1α	IFN γ	CD-16	*p-Value
Pree-Vaccination	44	5.786 ± 0.915	113.356 ± 26.052	2.024 ± 0.228	31.034 ± 7.094	0.000	0.657
Post-Vaccination	44	6.581 ± 2.472	132.992 ± 29.298	1.027 ± 0.108	22.777 ± 2.840	0.000	0.657
**p-Value		0.517	0.333	0.003	0.916

Correlations Spearman's rho
Immunologic Markers	Correlations Spearman's rho	IL15	MIP 1α	IFNγ	CD16
IL 15	Correlation Coefficient	1.000	0.153	0.251	-0.148
	Sig. (2-tailed)		0.322	0.100	0.338
	N	44	44	44	44
MIP 1α	Correlation Coefficient		1.000	0.162	0.047
	Sig. (2-tailed)			0.293	0.759
	N		44	44	44
IFNγ	Correlation Coefficient			1.000	-0.087
	Sig. (2-tailed)				0.575
	N			44	44

Subjects	Comorbid						Pearson Chi-square	Odds Ratio	p-Value
	<4		>= 4		Total
	n	%	n	%	n	%
Sex
Male	6	13,6	1	2,3	7	15,9	1.82	0,167	0,177
Female	36	81,8	1	2,3	37	84,1
Total	42	95,5	2	4,5	44	100
Ages (year)
60-69	25	56,8	1	2,3	26	59,1	0,188	-	0,910
70-79	16	36,4	1	2,3	17	38,6
< 80	1	2,3	0	0,0	1	2,3
Total	42	95,5	2	4,5	44	100

Treatments	N	Mean±SD (HI Unit)			*p-value
Treatments	N	A/Singapore – EFTHA 1835	A/Singapore – EFTHAS 1828	B/Maryland – EFTTHAC 1822	*p-value
Pree Vaccination	44	204.55±27.15	145.00±27.7	41.82±4,51	0.000	0,000
Post Vaccination	44	1044.09±191.22	463.64±71.02	289.55±49.53	0.000	0,000
**p-value		0.000	0.000	0.000

CorrelationsSpearman's rho
Vaccine Types		IL15	MIP 1α	IFN γ	CD16
A/Singapore – EFTHA 1835	Correlation Coefficient	0.197	0.178	-0.02	0.221
	Sig. (2-tailed)	0.199	0.249	0.899	0.149
	N	44	44	44	44
A/Singapore – EFTHAS 1828	Correlation Coefficient	-0.054	-0.014	-0.265	0.373^*
	Sig. (2-tailed)	0.73	0.927	0.082	0.013
	N	44	44	44	44
B/Maryland – EFTTHAC 1822	Correlation Coefficient	0.112	-0.189	0.109	0.323^*
	Sig. (2-tailed)	0.469	0.218	0.483	0.033
	N	44	44	44	44