The COVID-19 pandemic has generated a great impact worldwide both on the population health but also on an economic and social level. In this health emergency, a key element has been and still is the need for information, which has become a daily concern for many people. Social media represent powerful tools for searching and gathering health-related information, thus becoming a new place where health authorities need to be present to disseminate information of preventive measures like vaccines against COVID-19, as well as try to block information against these public health measures. The main goal of this study was to analyze the role that healthcare professionals have in Twitter to support the campaign of public institutions on vaccination against COVID-19. To address this study, an analysis of the messages sent on Twitter containing the hashtag #yomevacuno, between 12 December 2020 was developed using the NodeXL software (Social Media Research Foundation, Redwood, CA, USA), focusing on content analysis of tweets and users' accounts to identify healthcare professionals. The results show that healthcare professionals represent only 11.38% of users, being responsible for 6.35% of impressions generated by the network #yomevacuno. We can observe that traffic information generated by healthcare professionals is not significant in comparison with institutions (p = 0.633), but it is compared to common users (p = 0.0014). The most active healthcare professionals were pharmacists (40.17%), nurses (27.17%), and physicians (12.14%). Their activity (90.43% of messages) was mainly focused on sharing messages generated by other users' accounts. From original content generated by healthcare professionals, only 78.95% had a favorable storytelling on the vaccine, but without sharing information about vaccines or vaccination. As a conclusion for this study, the participation of healthcare professionals in the dissemination and generation of information within the #yomevacuno communication strategy, led by the Spanish Ministry of Health, has been scarce. We emphasize the need to enhance communication skills in social networks to support public health campaigns through these increasingly important social media.