Born and raised in the province of Bergamo, Astori started playing football with local team Pontisola before joining Milan in 2001. He spent five years in the club’s youth system. Astori later reflected on his development, saying: “After a year of courtship, of auditions, I came to wear the AC Milan shirt with the very young regional players. Along with twelve, thirteen new players. There was always a lot of change. I’m from San Giovanni Bianco, so it was already an adventure. But the greatest memories are precisely due to transfers with teammates, making the return from training. They were beautiful years, lived positively, for a boy there was no better experience. Honestly, it is difficult to get out of balance, because in fifty professional clubs there are twenty boys for each team in the youth sector, so every word is illusory. I have always believed in it and hoped for it, growing from year to year until it becomes one.”

Astori played 4 games for Italy’s under-18 team, beginning in 2004. He did not receive any further call-up at higher levels of youth international football.

Astori made his official debut in the top-flight during an away game against Siena on 14 September 2008, coming on as a 53rd minutes substitute, in a 2–0 loss. Since making his debut for the club, he spent most of the season on the substitute bench throughout the season. But Astori made six starts for Cagliari, all of them were centre–back position. At the end of the 2008–09 season, Astori made eleven appearances for the side.

In June 2009, the club renewed their ownership with A.C. Milan to keep Astori at Cagliari for another year. At the start of the 2009–10 season, he replaced Diego López and quickly became a first team regular, playing in the centre–back position. Astori then helped the club kept three consecutive clean sheets between 28 October 2009 and 8 November 2009. He then scored his first goal for the club in a 2–2 draw against Fiorentina on 31 January 2010. A month later on 28 February 2010, Astori scored his second goal in a 2–1 loss against Chievo. On 3 April 2010, Astori scored an own goal against A.C. Milan to allow Milan scored the third goal in the match which Cagliari lost 3–2. Despite missing three matches during the 2009–10 season, Astori made thirty–five appearances and scoring two times in all competitions.

Over the summer, rumours spread when A.C. Milan was keen to sign Astori but nothing happened. Despite the rumours, Astori was keen to rejoin A.C. Milan in the summer. The club renewed their ownership with A.C. Milan to keep Astori at Cagliari for another year in June 2010. At the start of the 2010–11 season, he helped the side kept three clean sheets in the first five league matches. Since the start of the 2010–11 season, Astori continued to regain his first team place in the centre–back position, forming a partnership with Michele Canini. He also started in every matches until missing one match for picking up five yellow cards and served a suspension against Catania on 12 December 2010. After serving a one match suspension, Astori returned to the starting line–up against Cesena on 18 December 2010, starting the whole game, in a 1–0 loss. Following his return, he continued to start in the centre–back position for the next twelve matches. This last until Astori was suspended for the second time for picking another five yellow cards and served a suspension against Napoli on 20 March 2011. After serving a one match suspension, Astori returned to the starting line–up against Genoa on 3 April 2011 and helped the side kept a clean sheet, in a 1–0 win. In a follow–up matcha against Brescia, he set up the club’s opening goal of the game, in a 1–0 win. On the last game of the season against Parma, Astori set up the equalising goal, in a 1–1 draw to end Cagliari’s four match losing streak. At the end of the 2010–11 season, he made thirty–six appearances in all competitions.

Astori received his first call-up for the senior team under newly appointed coach Cesare Prandelli who named him in the squad for a friendly against Ivory Coast, to be played on 10 August 2010; however, he was left on the bench as an unused substitute. He made his full debut for Italy on 29 March 2011, in a 2–0 friendly win against Ukraine in Kiev. He came on as a 17th-minute substitute for the injured Giorgio Chiellini but was sent off with 15 minutes remaining in the second half after receiving two yellow cards. A year later, Astori was named in the provisional 32-man squad ahead of the UEFA Euro 2012. However, he was subsequently cut from the squad on 28 May 2012. But following Andrea Barzagli’s injury, there were speculation on whether Astori would be Barzagli’s replacement. Eventually, Barzagli was kept in the squad and Astori stayed with Italy’s training camp until 9 June 2012. Two months later, Astori was called up to the Italy and started the whole game, in a 2–1 loss against England on 15 August 2012.

On 22 June 2011, the club bought the residual 50% registration rights from Milan for another €3.5 million. (pre-agreed €1 million tagged in 2008 and additional €2.5 million) At the start of the 2011–12 season, Astori started in the first eight league matches, playing in the centre–back position, continuing to partner with Canini. However, he suffered a fractured foot in a match against Napoli on 23 October 2011 following an intervention by Ezequiel Lavezzi and have to come off on the 21 minutes in a 0–0 draw. His injury resulted in his being out for 3 months. After the match, Astori revealed that Lavezzi personally apologised for injuring him. It wasn’t until on 8 January 2012 when he returned to the starting line–up, in a 3–0 win against Genoa. Since returning from injury, Astori regained his first team place, playing in the centre–back position, for the next twelve matches. But he missed one match for picking up five yellow cards and served a suspension against Atalanta on 1 April 2012. It wasn’t until on 7 April 2012 when Astori returned to the starting line–up against Inter Milan and scored his first goal of the season, in a 2–2 draw. He later regained his first team place, playing in the centre–back position, for the rest of the season. At the end of the 2011–12 season, Astori made twenty–nine appearances and scoring once in all competitions.

On 9 July 2012, Astori revealed he turned down a move to Russian side Spartak Moscow worth a €15 million (£11.9 million) fee after agreement between two clubs. Instead, Astori signed a contract with Cagliari that keep him until 2016. Astori explained turning down a move to Spartak Moscow because he wanted to stay at Cagliari and achieve his goals with the club. At the start of the 2012–13 season, Astori appeared in the first two league matches for Cagliari before being dropped from the squad for the next two league matches, due to sustaining a muscle injury while at international duty. He then returned to the starting line–up, starting the whole game, in a 2–0 loss against AC Milan on 26 September 2012. Astori then helped the club kept three clean sheets between 7 October 2012 and 28 October 2012. But he was suspended on two occasions, including his sending off for a second bookable offence, in a 3–1 los against Juventus on 21 December 2012. In the January transfer window, Astori was linked with a move to Premier League side Southampton, but the move fell through and he stayed at the club. Astori was once again sent–off for the second time for a second bookable offence, in a 1–1 draw against AC Milan on 10 February 2013. He then captained for Cagliari for the first time in his career against Napoli on 21 April 2013, as they lost 3–2. Despite being sidelined throughout the 2012–13 season, Astori continued to regain his first team place, playing in the centre–back position. He went on to make thirty–three appearances in all competitions.

Ahead of the 2013–14 season, Astori was linked a move away from Cagliari with Napoli interested in signing him. But he ended up staying at the club. As a result, Astori was dropped to the substitute bench amid to his future at Cagliari. It wasn’t until on 15 September 2013 when he made his first league appearance of the season, starting the whole game, in a 1–1 draw against Fiorentina. Since the start of the 2013–14 season, Astori continued to regain his first team place, playing in the centre–back position. He then captained the side for the first time in the 2013–14 season against Parma on 15 December 2013 and helped the side kept a clean sheet, in a 0–0 draw. In a follow–up match against Napoli, Astori set up the opening goal of the game, in a 1–1 draw. After missing a match against Livorno on 16 February 2014 due to “muscle ache in the rectus femoris of the left leg”, he returned to the starting line–up and captained for Cagliari in the next two matches, earning four points for the side. But after serving a one match suspension for picking another five yellow cards and served a suspension against Lazio on 16 March 2014, Astori returned to the starting line–up and captained Cagliari in the next two matches, earning three points for the side. Having captained the total of six times in the 2013–14 season, Cagliari News 24 made an assessment of Astori’s performance, as he alternated good matches with less good performances. As the season was concluded, Astori went on to make thirty–five appearances in all competitions.

Astori was a member of the 23-man Italy squad that took part at the 2013 FIFA Confederations Cup under Prandelli; Italy reached the semi-finals, only to suffer a penalty shoot-out defeat to Spain following a 0–0 draw after extra time, on 27 June 2013. On 30 June 2013, Astori scored the opener against Uruguay in the third place match after knocking in the rebound from Alessandro Diamanti’s free kick, as Italy won 3–2 on penalties to win the bronze-medal, following a 2–2 draw after extra time; this was his only international goal, and he became the first Cagliari player to score for Italy since Luigi Riva had done so 40 years earlier, in a 2–0 victory over Switzerland on 20 October 1973. A year later in September 2014, Astori was called up to the national team and earned his eighth international cap, in a 1–1 draw against Netherlands. Three years later on 5 September 2014, he played his last international match for Italy, starting the whole game and helped the national side kept a clean sheet, in a 1–0 victory over Israel. In total, Astori made 14 senior appearances for Italy between 2011 and 2017.

On 24 July 2014, Cagliari announced Astori’s move to Roma on a season-long loan deal. The transfer was settled in €2 million and Roma held the option to make it permanent at the end of the season for another €5 million. It came after when Astori renewed his contract with Cagliari a day before the Roma transfer.

Astori made his Roma debut in the opening game of the season against Fiorentina and helped the side kept a clean sheet, in a 2–0 win. Two weeks later on 17 September 2014, he made his UEFA Champions League debut, starting the whole game, in a 5–1 win against CSKA Moscow. However, after the match, Astori suffered a knee injury that kept him out for three weeks. It wasn’t until on 18 October 2014 when he returned to the starting line–up against ChievoVerona and helped Roma kept a clean sheet by winning 3–0. Astori then followed up by keeping two clean sheets in the next two matches against Sampdoria and Cesena. However, during a 2–0 loss against Cesena, he suffered a hamstring injury that saw him substituted and was sidelined for two weeks. It wasn’t until on 22 November 2014 when Astori returned to the starting line–up, in a 2–1 win against Atalanta. Two months later on 6 January 2015, he scored his first goal for the club, scoring the only goal of the game, in a victory over Udinese. Throughout the 2014–15 season, Astori found himself in and out of the starting line–up for the side, as he continued to fight for his place over the centre–back position. Having helped Roma finished second place to qualify for the UEFA Champions League next season, Astori made 30 total appearances during his time at the Stadio Olimpico, and scoring once in all competitions.

Following this, it was speculated on Astori’s future on whether Roma would sign him on a permanent basis. But it was reported on 23 June 2015 that Roma has decided against signing Astori on a permanent basis.

On 4 August 2015, Astori signed on loan for Fiorentina until the end of the season with an obligation to buy. Upon joining the club, he was given a number thirteen shirt ahead of the 2015–16 season.

Astori’s first game after signing for the club on a permanent basis came in the opening game of the season, starting the whole game, in a 2–1 loss against Juventus. This was followed up by keeping a clean sheet in the next three matches. However, he suffered a muscle injury that kept him out for two matches. It wasn’t until on 2 October 2016 when Astori returned to the starting line–up against Torino and started the match, as Fiorentina lost 2–1. Three weeks later on 26 October 2016, he scored his first goal for the club, in a 1–1 draw against Crotone. Following his return, Astori continued to regain his first team place, playing in the centre–back position for the next three months. This last until he missed one match for picking up five yellow cards and served a suspension against Pescara on 1 February 2017. After serving a one match suspension, Astori returned to the starting line–up, in a 4–0 loss against Roma on 7 February 2017. He then helped the club kept four consecutive clean sheets between 3 March 2017 and 2 April 2017. Astori then played an important role in the UEFA Europa League, setting up two goals for Fiorentina that saw the club progress to the knockout stage. However, they were eliminated in the knockout stage after losing 4–3 to Borussia Mönchengladbach in the UEFA Europa League Round of 32. He then scored his second goal of the season, in a 5–4 win against Inter Milan on 22 April 2017. Despite being suspended on two occasions later in the 2016–17 season, including a second bookable offence against Palermo on 30 April 2017, Astori went on to make forty appearances and scoring two times in all competitions.

At the beginning of 2017–18 season, Astori was appointed as the team captain following the departure of Gonzalo Rodríguez. He captained his first match at Fiorentina, starting the whole game, in a 3–0 loss against Inter Milan in the opening game of the season. Three weeks later on 10 September 2017, Astori scored his first goal of the season, in a 5–0 win against Hellas Verona. Since the start of the 2017–18 season, he continued to regain his first team place, playing in the centre–back position. Astori then helped the club’s defence kept four league consecutive clean sheets between 3 December 2017 and 22 December 2017. Since the start of the 2017–18 season, he started in every matches until he missed one match for picking up five yellow cards and served a suspension against Sampdoria on 21 January 2018. After serving a one match suspension, Astori returned to the starting line–up, in a 4–1 loss against Hellas Verona on 28 January 2018. On 25 February 2018, Astori made his last appearance for the club, playing the full match in a 1–0 home win over Chievo; six minutes into the match, Astori made a run forward, joining the attack, and passed the ball to Cristiano Biraghi, who scored the only goal with a long-range shot.

On 4 March 2018, Astori died in his sleep while staying in a hotel in Udine prior to Fiorentina’s match against Udinese, proven to be caused by cardiac arrest determined from an autopsy conducted two days later. A club statement said “Fiorentina are profoundly shaken and forced to announce that their captain Davide Astori has died.” His former Cagliari teammate and coach Diego López and former Fiorentina teammate Carlos Sánchez collapsed to the ground after hearing the news of his death, however they both recovered shortly after. All Serie A and Serie B matches scheduled for 4 March were postponed. His death had caused shock across the footballing world, and many coaches and players paid their respects. UEFA later announced on Twitter that a minute’s silence was to be observed before every European match scheduled for that week. Cagliari and Fiorentina both retired the number 13 jersey worn by Astori in his honour (Astori wore this number as it was the number that his idol, Alessandro Nesta, had also worn throughout his career). He was buried at San Pellegrino Terme cemetery on 8 March 2018. Fiorentina’s next home game against Benevento, on 11 March, was paused for 60 seconds at the 13 minute mark in his honour, while children from both teams entered wearing Astori’s jersey to pay tribute to him; Fiorentina won the match 1–0, with the only goal coming from Vitor Hugo, who celebrated after scoring by saluting a T-shirt bearing Astori’s image.

In April 2019 it was reported that Astori’s contract would be renewed for life with all the money going to his wife and child, but it was later revealed to be a hoax.

In May 2019, during the 2018 ceremony for new members being inducted into the Italian Football Hall of Fame, a Fair Play Award in memory of Astori was also awarded for the first time ever; the recipient of the award was Igor Trocchia.